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1. 0 118 Alleles How are they designated 0 0 2 eee ee eee eee 118 Matched Reaction Pairs MRPs How does LTI identify 120 MRP Table How does LTI populate the o oooooooooooooooo o 122 MRP Table How are Allelel and Allele2 grouped in the 124 All Alleles Table How is the generated oooooooooooomomomooo 126 Type Subtype Allele Assignments How are made 127 Reaction Grid How is the sorted 2 2 eee eee eee 128 NMDP Codes How can I automatically update l 2 2 2 ee eee 130 LTI database directory Why is the growing excessively 131 How do I update an LTI database with a non default name 132 How do I install MSDE to a non default folder ooooo ooooo o 133 How can I import LABType Classic analyses into HLATools 134 Version 1 1 HLATools User Manual 4 of 172 OLI UD 0127 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Appendix C Network Installation of HLATools Glossary Peer to Peer Instaulaons 4 4 0 44 0044 4044 1994 4909495 Raco 137 Creating a Database on a Remote Server 2 0 0 0 eee eee eee 138 Connecting to a Remote LTI Database 0 eee ee 140 Editing the Authorized Users List from a Remote Server 148 Linking to another Server using SQL Authentication 149 Setting up HLATo
2. o um a DU v a uw a D m a o va oO va amp g uw un D oO a a 1 Sero1 2 and Bw 1 2 or Cw1 Cw2 serological equivalent and for B locus or C locus typing Bw or Cw allotypes for each detected allele To transfer the cell contents for an allele from the table to the corresponding assignment fields double click on one of the Sero or Bw or Cw cells for that allele Use the Restore key to revert to the orig inal computer assignments 2 DNA allele assignment fields for Allele 1 and Allele 2 3 Serological and Bw or Cw serology and where appropriate Bw or Cw allotype assignment fields Serological equivalents displayed here will be included in the patient and sample reports when the sample has been reviewed and accepted The serological equivalents and Bw4 Bw6 or Cw allotype entries are editable allowing the reviewer to override the computer generated assignments To edit these values Enter new value s into the Serological or Bw or Cw edit fields e Click Accept to commit the changes to the database The emended serological equivalents will appear in the reports but the original values are maintained in the table The DNA assignments now appear in bold face type to indicate that the allele assignments have been reviewed and Accepted Allele assignments that have been Accepted are designated Corrected Typings in the reports When the assignments for a sample have been accepted
3. The HLATools Patient Explorer associates multiple sample results collected from among one or more batches with a single patient These results are assembled and collated on a patient by patient basis and presented for review here Reports of the collected sample results can be printed out in Patient Summary reports via the Lambda Reporter Important Note Patient typings that have been accepted are designated corrected in some reports Overview The patient information displayed here can be entered and edited in the LTI Patient Information Subview p 57 1 Patient ID see Patient Information Subview p 57 for details on using the Patient ID to associate multiple sam ples with a single patient All assignments for patients with samples associated to their Patient IDs in this and other active batches are listed here Sample results for patients in other active batches are not displayed here unless the patients are also present in the selected batch 2 Date of Birth non editable field to change the date display format see Changing Date Formats in HLATools p 107 Last Name non editable field Comments patient specific comments viewable via a tool tip expands closes the subtable of sample information associated with each patient DNA ape e ConsensusAlleles 1 amp 2 the NMDP code for sample allele assignments If the patient is homozygous at this allele only the first cell conta
4. Reaction Grid How is the sorted The following heuristic method is employed to sort alleles in the Reaction Gird to allow the user to examine how similar they are to the Sample Reaction Pattern The allele sorting is driven by two user specified alleles referred to below as First Allele and Second Allele which are entered in the text search fields at the top of the grid Figure 5 16 Five different types of scores are assigned to each allele A1 A2 PP FN and TP Typically these scores are not displayed in the Reaction Grid 1 Al scores are assigned to the sample reaction and to the alleles as follows e The Sample Reaction is always scored 1 e Alleles that share fewer than 75 positives with the First Allele are scored 0 e Alleles that share more than 75 positives with the First Allele are scored 5 unless e they also share the first four digits family and subtype designations with the First Allele in which case they are scored 3 There will always be at least one 3 If an allele designation includes an NMDP or local am biguity code the comparison is made on the basis of the numeric allele designations defined by the alphabetic code 2 A2 scores are assigned to the sample reaction and to the remaining alleles after the positive reactions have been clustered to the left of the grid The left part of the gird is defined as the string of consecutive positives in Allele 1 whe
5. Assignment Patient 1 Patient ID this is distinct from the Sample ID and is often different from the patient s name for confidentiality reasons A Patient ID is associated with all samples belonging to the patient Many samples can be associated with a single patient but only one patient can be associated with a sample Clearing the Patient ID then pressing Match removes all entries from the subview 2 Last First Middle last and first names are included on results reports 3 DOB the DOB combo box accesses a calendar utility that defaults to 01 01 1901 however the field also accepts direct entry of a date which is parsed by the calendar utility This makes it easy to jump to any desired date 4 Text field comments entered in this text field are included in the Patient Reports that can be printed out from the Lambda Reporter 5 Match if you enter a preexisting Patient ID in the Patient ID field Match associates the current sample with that patient the preexisting patient information will be displayed here as well Executing the Match function also Version 1 1 57 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf enables the Save function After using Match to associate a sample with a patient be sure to Save the newly estab lished association To dissociate a sample from a patient clear the Patient ID field then Match and Save 6 Save commits any edits to the database and locks the subvie
6. 0 0 0 e ee eee eee 107 Changing Batch Results Column Order 0000000000 108 Navigating the LTI Interface 22 c2eiceri caddie eee bacdkarebevauherteavans 109 Navigating Between Windows Using the Keyboard 109 Accessing LTI Functions via Hot Keys 0 0 00 ee eee eee 109 Changing Date Formats in HLATools 0 000 0 cee eee eee 109 Editing the Assignment Subview Comments LiSt oooooooocoooooo o 109 Specifying the Data Transfer File Export Type 0 0 0 0000 110 Specifying the Character Encoding Type 0 e cece eee eee eee 111 Appendix B HLATools FAQ Frequently Asked QuestOnS 2 2edweeeeicdeseensedsesevenGhseteeeaus 113 How can I tell if HLATools is using the most recent LABType products 113 Why can t I find the Admin Diagnostics tool 2 2 0 0 eee ee eee eee 113 Update Database Why can t I run coco 114 LTI database does not exist Why do I get a message that 114 Remote LTI database Why can t I connect to using Windows XP 114 Open a port though the XP firewall How dol ooooo oooooooo o 114 Local security policy on XP How do I modify 020008 116 Clean Database Why do I get a BCP error 0 0 0 0 00 0 e eee 116 Why do I have to log into the database when I try to print some reports 118 Windows Authentication Mode Why can t Isetup
7. Configuration LabType View Update LabScreen Connections Database Attach Detach Backup Restore View Update Local Create Merge System Diagnostics User Management view Update Filter NMDP o Serological Update Products gt View The HLATools Product Information table lists all the LABType product lots that are included with the current release of the software Several columns in the table contain important editable default values that govern how batch data associated with a given product lot are handled by LTI and which type of data LTI uses Figure 7 2 Product Information Controls Tools Refresh Cutoff Sensitivity The controls above the HLATools Product Information table have the following functions e Tools gt Global Settings menu selection accesses the Global Values dialog where you can set the Minimum Beat Count and Minimum Positive Control values for all products at once e Tools gt Manage menu selection sorts the products in the Product Information table by descending order in which they appear in the Lambda Explorer Product Lot Selection pull down menu Figure 4 4 You can edit the values in the Visible column to change their order of appearance in the menu 72 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Refresh fetches values currently stored in the database and
8. Figure 8 8 Different sets of appropriate search operands are available for filtering on each row Figure 8 8 Advanced Search Filters FE Enter filter criteria for Patient ID i x Operator Operand And conditions El Greater than gt Sample B4 Equals to i Add a condition Does not equal to Less than Less than or equal to Greater than Greater than or equal to Like Sample B 7 Matches Regular Expression Sample B8 Delete Condition Cancel l Patient ID gt Sample B4 AND Patient ID lt Sample B 86 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 8 You can preview the reports for the selected samples in the specified format by clicking on Preview in the Report selector pane see bottom of Figure 8 4 Figure 8 9 shows part of a Patient report Figure 8 9 Patient Report for One Sample Your Lab Your Street Your Town CA 90000 Your Director Tel 800 555 1212 Fax 818 555 1213 E mail you yourlab com Renewed By LABType SSO Class Data i cal Sample ID 2436135 Catalog4ot FASSO1B_006 Rev 03 Locus B Batch Name 123 104B_L6 Analysts Date 5 5 2006 Tested By Test Date 11 2 2404 File Path C LuminexFilesd23_104B_L6_ID1473 csv Batch Analysts Comment Patient ID 2436135 Gender ABO Rix Patient Name Race Comments Corrected Typing Computer Assignment B 44RDU B 57Gw Generic Allele Descriptions B 44RADU gt 0514 17 B 57GW gt 010608
9. Interface Nomenclature e The term View is used instead of window or tab e A tabbed view is termed a View or Subview e A Panel or a Pane is a demarcated region or area in a View or Subview 12 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf e A Batch is a collection of Samples and appears as a folder in the Batch Samples file tree Note that the contents of a batch are termed samples and not patients This distinction is made because multiple samples may be associ ated with a single patient and because the confidentiality policies of some institutions require that the patient sam ple relationship not be openly divulged The treatment of patients with multiple samples is addressed in Chapter 6 Patient Explorer e The term Bead is often used synonymously but imprecisely with Probe since a LABType bead can be assumed to carry only one type of probe Technical Support For technical support or to report software problems contact your One Lambda representative e From the United States call 800 822 8824 e In the greater Los Angeles area call 818 702 0042 e Contact us by e mail at softwaresupport OneLambda com Version 1 1 13 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf This page intentionally left blank Chapter2 Installation and Log On This chapter addresses installation of HLATools software on a single user standalone workstation Installation
10. OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Chapter 3 LABType Home a LABType Home When you launch HLATools M the icon at the top of the list bar is the LABType Home icon shown at the right This icon accesses an HTML page with links to One Lambda product information in PDF format for LABType SSO and Micro SSP product lines The documents are included in the software distribution and reside on your local computer or a network server thus allowing you to access the information without going onto the Internet This HTML page contains links to four types of SSO reference documentation Product Inserts Product insert sheets for each typing test are provided in the principal European languages required by the EU IVD Directive A Product Insert contains the protocol that must be followed to carry out a LABType SSO typing test Patient Worksheets Patient Worksheets are used to record the reaction results for each bead in a patient sample Each worksheet contains the locus allele groups and their serological equivalents and positive cutoff values for each bead The worksheets are intended for use by laboratory technicians and HLA typing experts Bead Probe Information These sheets include each bead s probe recognition site start and end points of various locations in HLA protein sequence alignments that are recognized by the specific probe allele specificities for the probe the One Lambda Version 1 1 27 of 172
11. 2 Access the SQL Server 2000 Service Pack 3a download page at http www microsoft com sql down loads 2000 sp3 asp 3 From the download drop down list select the language version you wish to download and press GO This will take you to the download page for the specific language At the bottom of this page are several download links 4 Choose the file of the format lt lang gt _sql2kdesksp3 exe where lt lang gt is the abbreviation for the language you have selected Download this file to your desktop 6 Click on the file This will open an Installation Folder dialog Accept the suggestion for the default installation folder C sql2ksp3 unless you want to save the file to a different location The file will be unpacked in this folder 7 From the desktop select Start gt Programs gt Accessories gt Command Prompt Navigate to the installation folder mentioned above 9 Execute the command Version 1 1 115 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf setup upgradesp sqlrun INSTANCENAME ONELAMBDA DISABLENETWORKPROTOCOLS 1 To update the default instance of MSDE for other applications that may use it other than LTT 1 Repeat steps 7 through 8 above 2 Execute the command setup upgradesp sqlrun DISABLENETWORKPROTOCOLS 1 For more information on SP3 SP3a see article 819334 on the Microsoft Support website Why do have to log into the database when try to print some re
12. N or low expression L alleles are not expressed on the cell surface or are expressed to a very low degree They are desig nated by an N or L suffix Phosphate Buffered Saline a non toxic saline solution used as a diluent in sample preparation PCR Sequence Specific Primer in PCR SSP oligonucleotide primers are designed to obtain amplification of specific alleles or groups of alleles The typing method is based on the premise that a completely matched primer will be more efficiently used in the PCR amplification than a primer with one or more mismatches The One Lambda Micro SSP product employs PCR SSP analysis PCR SSP has a number of advantages over other PCR based typing methods 1 High degree of resolution with each primer pair defining two linked cis located poly morphic sites which facilitates the typing of heterozygous individual 2 Results are easy to interpret The inefficiency of the Taq polymerase in extending mismatched primers is a more precise reaction than hybridization with ASOs HLATools User Manual 163 of 172 HLATools_IVDManual_v1 1_ Rev 0 pdf Positive Control Positive Reaction Primary Data Files mdf Recognition Site 4 The post amplification analysis is more rapid and simpler than other PCR based methods because the typing specificity is part of the PCR 5 Inexpensive and versatile it can be used for all Class II loci 6 For higher resolution a second analysis
13. 2 From the desktop menu select File gt New gt Folder 3 Name the folder exactly LuminexFiles and press Enter Use this folder to store your Luminex LABScan 100 csv files If you have installed the HLATools software somewhere other than in your C drive you can use the built in browse function to locate csv files for analysis For instructions on how to access Luminex output files directly from the computer that controls your Luminex analyzer see Sharing Luminex Files on a Network p 150 Running the Clean Database Utility This utility creates the database structure required by LTI and other HLA Tools applications The database files are located in the folder C Program Files Microsoft SQL Server MSSQL QONELAMBDA Warning If you are installing HLATools for the first time you must run the Clean Database utility before launching any of the HLATools applications You must have administrative permissions in order to run Clean Database If you attempt to use the Lambda Explorer to import files or run LTI without previously creating a database both applications will generate a warning message and quit If you are updating HLATools you probably want to preserve your existing analysis results To determine if you should run Clean Database or Update Database see Updating HLATools p 23 To run Clean Database 1 Select Start gt Programs gt One Lambda gt HLATools Configuration Suite gt Clean Update 18 of 172 6 6
14. 9 The Filter panel also contains a Report Summary subpanel Figure 8 10 that can be toggled via the Hide Show Report Summary checkbox in the Sections tab of the Search subpanel Figure 8 11 You must first click the Edit button to enable the checkboxes in the Sections tab Figure 8 10 Report Summary Subpanel Report Summary User supplied report specific comments entered at the time of report generation M Show Report Comments on Report Generate the report by selecting a patient The report will include only the samples corresponding to the analysis from the patient selected Checking Show Report Comment on Report includes the report specific user comments in the report Figure 8 15 The lower text field contains general instructions about the current type of report being generated This text does not appear in the report Version 1 1 87 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Creating a Custom Report Template The easiest way to illustrate how to create a custom report template is by modifying a stock template Figure 8 11 Sample Report Default Sections Search Search Sections Paper Size Logo M Lab Information Y Patient Information Rixn Pattern PC Beads Graph amina Messages Hide Report Summary V New Page After Reviewed Information Edit Save Cancel In the Lambda Reporter e Select the report category Sample Reports at the bottom of the Report
15. Cutoff val ues for beads can be changed locally on a sample by sample basis in the Reaction Pattern Histogram in the Assign ments view Figure 5 8 10 Globally Changed Cutoff to change the cutoff value for the bead globally e clear the Locked check box 17 e click on the location for the new value for the cutoff this automatically relocks the view To restore a globally changed cutoff to the default value e clear the Locked check box e click on the blue default Cutoff line the global change line will disappear 11 Sample Specific Change Indicator when a bead cutoff has been adjusted in the Reaction Pattern Histogram for a specific sample this will be indicated by an arrowhead that points in the direction of the previous setting In Figure 5 4 bead 59 has been changed globally as indicated by the repositioned global cutoff line and then locally in sample 2436212 as indicated by the upward pointing arrowhead 12 Tool tip mousing over a column displays the sample ID number and the normalized bead reading double click ing on the column opens the Assignment view for the sample 13 False Reaction Indicator shows the number of times that a bead has been used in an allele assignment involving a single change in reaction status for a given bead i e a Close Reaction False Positives are bright red and False Negatives are dark green Changes in cutoff values are immediately reflected in this chart 14 Y axis number of
16. DPB1 5901 3 FN Allelet items Allele1 DPB1 4501 2 items Allele2 FalseReactions DPB1 5701 42 FP Allele1 DPB1 6801 1 item Allele2 FalseReactions lt Collapse DPB1 9101 43 FN Apypads swcyasasop addygns adAy sajeg iiy paynew Changing Batch Results Column Order You can modify the Batch Results Grid table so that its columns appear in a different order Figure A 2 Rearranging Table Column Order BatchResultsGrid Allele 1 Allele 2 Comment PatientID Lek Relati Famil CR MRP Por isos a 0801 waai T Fah ENT rer aos f ELLE iZ Wwe _ rer aos IO O CL Phones Ll ura m2 Gorse orerazma DPBTIIN NoFabeReactons _ JomBrewser JE 14 2 _ For example as shown in Figure A 2 you can rearrange the columns so that the Patient IDs appear directly to the right of the Sample IDs by dragging the Patient ID header from its present location and dropping it next to the Sample ID header You can save the modified column order to that it is persistent from session to session See the discussion to Figure 5 3 for details on adding or deleting columns from this table 106 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Navigating the LTI Interface Navigating Between Windows Using the Keyboard Many of the functions and navigation features in LTI can be accessed using the hot key functionality familiar to users of Microsoft Windows To navigat
17. Generic Allele Descriptions No False Reactions 1 Matched Reaction Pair DRB1 0103 amp DRB1 15H5 gt DRB1 0103 amp DRB1 150101 DRB1 150103 DRB1 150105 DRB1 1513 DRB1 1516 4 Close Reactions 15 FP 25 FP 42 FP 62 FP O DRB1 0103 DRB1 15HS gt 01 3 16 Accepted gt workshop2 2006 05 08 16 01 04 Reviewed gt wwhite 2006 05 11 09 57 17 90 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 8 15 Non Default Sections of a Sample Report 510 2006 Page 1 of 1 Reviewed By LABType SSO Class Il Data Reviewed Date User supplied comments entered at the time of report generation 10 Sample ID Sample5 D Analyzed workshop 200605 08 1601 04 Locus DRB1 Reviewer Catalogd ot_FAS550281_008_Revw 03 wnwhite200605 11 0957 17 Corrected Typing DRE1 D1BREY DRI DRBI 14 HG DR14 Warning Messages Low Bead Count 12 13 RxnAssignament 81111811818118188111111111111111111811111181111111111111111111111118 Figure 8 16 Sample Report Positive Control and Reaction Pattern Histogram LAMISIL IIA IDA e Specifying Report Paper Size The default report paper size is normally determined by the Windows OS language specification The report paper size has U S Letter dimensions when U S English is specified in the Regional and Language Options Windows control panel or A4 size when any other language is specified However the paper size local default can be overridden in
18. LTI then searches for two aways three aways and so forth Searches for False Reactions are initiated when the user changes values for bead cutoffs either globally in the Bead Analysis Panel Figure 5 4 or locally from within a sample s Reaction Pattern Histogram Figure 5 9 118 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure B 4 Construct observed Sample Reaction Pattern SRP by comparing the normalized bead readings to cutoff values to determine positive reactions For each of the n alleles recognized by the selected L4BType product combine the known reaction pattern of Allele with that of Allele Il Do the Yes Continue matching with next allele serving as Allele SRP Record Allele li Allele ll pair as Matched Reaction Pair lls No Yes Yes an Have all alleles had their turn as Allele combined rxn patterns match the Close Has this Allele No Reactions been combined with all Allele LTI Analysis Flowchart Increment Mismatch Number To identify False Reactions repeat matching procedure to find allele pairs that match observed SRP to within one bead reaction fewer or greater record Allele Allele ll pair and ID s of missing or extra bead s To identify Close Reactions toggle bead reactions one by one repeat matching procedure for False each modified SRP
19. Merge Diagnostics User Management Add SQL User Add Windows User Group Delete User 3 Once the Homer connection has been made the users for Homer s computer appear in the display field just as though you were observing the list while logged into that computer see Figure C 5 4 With access to the target server swordfish established it is possible to add and delete users on the target server remotely in the same way as one modifies the user list when logged in locally Linking to another Server using SQL Authentication As an alternative to using Windows network security it is also possible to use SQL authentication to control access between servers In SQL authentication access is granted by using a username password combination that is different from the Windows username password Important Note By default MSDE SQL Server 2000 installation sets the login authentication to Windows authenti cation mode The authentication mode can be changed after installation to SQL mode or mixed mode both SQL and Windows by using the SQL Server Enterprise Manager or by appropriately editing the registry on all servers which are to operate in a peer to peer configuration Changes in authentication mode should be undertaken only by a systems administrator 1 Add an SQL user to the another computer s user list This can be done using the User Management tool locally or remotely Select Add SQL User and enter a username pas
20. Merge Database siisuVstarar dra ee baka bau e sense tee eee ead 163 A ee Daw oot E ancaane eee nee e4 Oe eee ea ew ene ceo ee Re 163 NADIE oon fatetiee tak sis dai genes dais ooh wake 163 Negative COUMN0l 24 veh ranee eee E e ce eete nestor enenseees vee Ke 163 IMTS CORE A A NS SEE a POI RD Ade ea eae aro 164 NMDP COGS i440 badeodsbeenaeweebat ae ehhe eke nhac hariheeens iene 164 IN OPIN ANI Z ALOT 9 6 g3 5 8 Sey ws Mea a a re ETE a ow SE Ck RO Pe 164 Version 1 1 OLI UD 0127 062006 HLATools User Manual 5 of 172 HLATools_IVDManual_v1 1_ Rev 0 pdf Nucleotide Codes o ooooomoooomo eeeneeecaveeuveeaes 165 Null All l 22 insista as rr a ao he dete a 165 PBS 2 2eticnte cc a etek ei ie he oe iG ae eee a yale Le ae ke 165 PCR SS Piers Ge eS abe Whoa he GE AA Ade 165 Positive Control 166 Positive ReactiOn vimos e a atea abs 166 Primary Data Files mdf arras sia o a a 166 Recognition Site cvs rar a a 166 REEP ios e e a ob 167 Secondary antibody rra la eii ds 167 DO Os a okra tals GRR ea 6 wee Wale cht ed NE a aQeoe Ney adele ee eaeele wines 167 SUBSEA taa scl aries staat coke id 167 Umcode renga a goa ee eee Bee Be 167 Update Databases psi do aaa 167 Index Version 1 1 HLATools User Manual 6 of 172 OLI UD 0127 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Chapter 1 Introduction to One Lambda HLATools This manual is intended to serve as in depth discussion of the features of the One Lambda HLATool
21. OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf reference test number for the bead information and the number of true positives and negatives and any false positives and negatives observed during the reference test Each new revision of the Bead Probe Information sheet also includes any changes or additions to the allele specificity list for each bead Resolution Limitations Molecular genotyping is capable of much finer resolution of alleles than serological testing This sheet contains a listing of the alleles detected by LABType SSO testing that exhibit the same combined reaction patterns at the serological level 28 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Chapter 4 Lambda Explorer Lambda Explorer The HLATools Lambda Explorer module is the tool used to import LABScan lt SuperScript gt 100 output files into the HLATools database When you first access the HLATools Lambda Explorer view the batch files tree displays all of the Luminex files that have been loaded into the HLATools database or which are currently available for loading Warning Before you use HLATools for the first time you must create a database structure by running the Clean Database utility See Running the Clean Database Utility p 18 for details Accessing Luminex csv Files When first launched Lambda Explorer looks for a folder called C LuminexFiles This file is the default repository for t
22. Only one value for MinPCData is speci fied even with products that have two positive controls such as lots for B locus analysis The value for MinPCData can also be changed globally using the Tools gt Global Settings menu option Visible indicates the priority ranging from 0 to 10 in which a product appears in the Product Select drop down menu when importing a sample batch into the LTI database from within the Lambda Explorer e 0 removes the product from the suggestions in the drop down menu see Figure 4 3 Selecting a File for Im port To reduce the clutter in the drop down menu that would be caused by including out of date products set this value to 0 However since you may need to review LABType analyses that were based on earlier prod uct lots it is recommended that you not delete out of date products from the Product Information Table e 10 is the highest priority and causes the product catalog lot to appear at the top of the dropdown list Other numbers from 1 to 9 can be assigned to force the order in which products appear Products with identical prior ity numbers are displayed in the dropdown list in the order they appear in this table Figure 7 4 Bead Data in the Product Information Subtable Bead Posctrl AbbreviatedSpecificity RecognitionSite ProbeID Cutoff CompleteSpecificity 13 13 Positive Control Exon 2 Positive Control Exon 2 OLR 4390 100 Positive Control 15 32 A 0252 A 300101 15 111 E
23. allele family equivalent of the serological antigen and the second pair the allelic subtype amino acid difference Examples would be B 4435 or B 5707 in Figure B 8 Groups of alleles with shared reaction patterns are designated with two digits and an NMDP or local ambiguity code For example B 44ANYV signifies that the allele may be B 440301 B 4426 B 4436 B 4438 or B 4439 Further digits or characters that are used to denote higher resolution alleles noncoding or synonymous polymorphisms intron digits and null or low levels of expression may also be shown in the Matched Reaction Pairs tabulation See the FAQ Alleles How are they designated p 116 Figure B 8 Matched Reaction Pair Tabulation Matched Reaction Pairs B 57XX4 B 570301 Allele1 Allele2 p BY44anyy BYS7xx4 B 4435 BYS7XX B 570302 B 4437 B 5707 BE44ANYY B 5707 B 4430 B 5707 B 4435 B 5707 ad xgnsfadAL sajaiiy Y paysqew Version 1 1 123 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf All Alleles Table How is the generated The All Alleles Recognized Allele Pairs Serological Equivalent table is created by matching all possible combina tions of the alleles in the Allele1 cell of the MRP table with all possible alleles in the Allele2 column cell For example in the first row of the MRP table B 44ANYV indicates that allelel may be B 440301 B 4426 B 4436 B 4438 or B 4439 5 possibilities and the matching allele2
24. e Install HLATools to a destination other than the C drive e Check available drive space The default option is the complete installation Click Next to proceed 11 The InstallWizard is now ready to complete the installation Version 1 1 17 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Click Install to proceed If for some reason you change you mind it is possible to abort the installation while it is in progress Click Finish to conclude the installation This completes the first part of HLATools software installation for a single workstation Before running HLATools you must create a folder where your Luminex files can reside and run the Clean Database utility to create an HLATools database structure See Creating the LuminexFiles Folder p 18 and Running the Clean Database Utility p 18 for further instructions Creating the LuminexFiles Folder When first launched the Lambda Explorer will look at the top level of the directory structure where HLATools has been installed for folders containing files in the csv format This is the format used for the Luminex LABScan lt SuperScript gt 100 output files that are the input files for the LTI analysis module In a standard instal lation to the C drive of your computer the Lambda Explorer will look for a folder called LuminexFiles As the first step of the installation create the folder for your csv files as follows 1 Select My Computer gt Local Disk C
25. gt View p 77 Updates code list from NMDP Research website or a local file For details see NMDP gt Update p 78 e Serological maintains allele serological equivalents list Displays equivalents list Exports list in plain csv or in Excel compatible csv format For details see Serological gt View p 78 Updates equivalents list from a website or a local file Comments Editor Constructs list of frequently used comments for user supplied sample annotation For details see Figure A 3 HLATools Comments Editor Tool 8 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Configuration General Configuration Lambda Explorer Default Directory specifies the pathway to the default folder containing Luminex 100 out put files accessed by the Lambda Explorer see Chapter 4 e Lab Info maintains laboratory and institutional information included in report headers For details see Entering Laboratory Information p 83 e Login Type sets level of HLATools login security see HLATools Security Modes p 149 LABType e Export Type specifies client specific data transfer file export types and enables client specific application fea tures e Character Encoding Type specifies the character encoding code page used in report generation LABScreen e Specifies the pathway to the folder containing the One Lambda LABScreen HLA analysis executable LAB Screen can be launched from HLATools explorer bar Fi
26. record Reactions Allele l Allele ll pair and Bead ID of toggled bead which yield MRPs for modified SRP Pair this Allele pattern with next Allele ll pattern Look for Close Reactions or False Reactions Have MRPs been detected or have searches for CRPs and False Reactions been completed Yes Record Allele l Allele ll pairs for MRPs plus bead IDs for Close Reactions and False Reactions Version 1 1 OLI UD 0157 062006 119 of 172 HLATools_ VDManual_v1 1_Rev 0 pdf MRP Table How does LTI populate the The process that LTI employs to populate the MRP table can be illustrated by referring to the Reaction Grid which is discussed in some detail in Chapter 5 see Reaction Grid p 58 e The x axis lists the beads in the assay including the control bead s In the default view the beads appear in as cending order from left to right Figure B 5 These are the bead numbers in the assays not the proprietary OLI bead identifiers Typically there are about 100 different kinds of probes in an assay and several hundred beads with each type of probe When the reaction grid for a sample is first accessed the topmost row of the table contains the sample reaction Rxn The beads with positive reactions are indicated by columns with light blue background Clicking on a row header cell causes all the reactions for that allele to be clustered to the left In Figure B 6 this has been done for the samp
27. the newly imported sam ple is also associated with the patient ID This allows all of a patient s samples to be reviewed together when you access the patient s analysis results in the Patient Explorer Chapter 6 Moreover analysis results for all samples associated with a patient can be printed out in the Patient Report which is described on page 95 Patient matches are not made if the samples are assigned trivial IDs e g 1 2 3 4 Version 1 1 33 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf This page intentionally left blank Chapter5 HLATools LABType iL Interactive LABT pe HLATools LabType Interactive LTI compares the results of the analysis of the sample data imported in the Luminex csv file to the reference information contained in the One Lambda Bead Probe Information and Resolution Limitations data sheets Analysis results are displayed in a number of interfaces that speed and greatly simplify user review Results are available at two levels of detail Batch Analysis and Sample Analysis Synopsis of LTI Batch Analysis Views The three principal views that address batch analysis tasks are shown schematically in Figure 5 1 A similar schematic for sample related views and subviews is shown in Figure 5 7 Figure 5 1 Batch Analysis Views HLATools Batch Sample Main Panel Explorer Bar Tree Three Tabbed Principal Views Batch Results Header and Table Bead Analysis QC Data and Sa
28. 0722 B 0730 B 0735 B sala IY payne adaygns add j Low Bead Count 2 Close Reactions 13 1 FN 21 FN Modified beads gt 1 Previous Accept Next 48 E Sample ID 24361 Date o O Birth 1 1 1901 Race Unk Assignment Patient 20 B 07A4MYN B 5001 sur y 1585017 wn D T 2 9 Mee gt tle 3 La t a tole kelley L ler E e e a a TNO LOCO P 00 M O Ode LO LO 000 O mrem rr is Ren Grid Raw Data Version 1 1 51 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 5 10 Allele Symbols in the Reaction Pattern Histogram DPB1 04UHY DPB1 8601 Selecting a cell in the Allele 1 or Allele 2 columns causes the positive beads in that allele to display their X or symbols beads in the other allele are indicated by the cutoff symbol Selecting both cells by clicking on the pointer at the left displays the symbols for all alleles Beads i e recognition sites present in both alleles are indicated by a diamond Matched Reaction Pairs Subview The Matched Reaction Pairs subview lists allele pairs that match the observed sample reaction pattern For the sake of brevity multiple alleles with identical reaction patterns are subsumed under an NMPD or local ambiguity code whenever possible This subview and its features are discussed above together with the Assignments View and the Sample Assign
29. 1_Rev 0 pdf Control Values View Minimum Bead Count 1 Minimum Bead Count the Y axis indicates the specified Minimum Bead Count for the sample or the observed bead count if the count for any individual bead falls below the specified minimum The Luminex 100 Analyzer continues to log beads until the threshold has been reached for every bead or until the well is exhausted The Minimum Bead Count can be set globally in the HLATools Product Information table Figure 7 3 The default min imum value is 100 2 Bead Count Symbol the actual bead count is indicated by a blue square 3 Tool tip mousing over a symbol displays the minimum bead count or the actual bead count if less than the mini mum Figure 5 6 Control Values Minimum Bead Count C Minimum Bead Count Minimum Bead Count Minimum Bead Count Sample81 SampleB2 SampleB3 SampleB4 SamplefS SampleBG SampleS Sample Bb Version 1 1 45 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Synopsis of Sample Analysis Views and Subviews There are four principal Views devoted to sample related tasks and analysis Assignments Bead Analysis Reaction Grid and Raw Data The Assignments view contains a main panel with five vertically tabbed subviews a panel on the right with two tabbed subviews for Sample Assignment allele typing and serology matching and Patient Infor mation patient demographic information the Pos
30. 8 HLATools Lambda Reporter Laboratory Report Header Information 0 0 0 cece eee eee eee ee 82 Entering Laboratory Information 0 0 c eee 83 The other fields are self explanatory and do not require elaboration 84 Using Lambda Reporter Filters 0 0 0 c eee eee eee 84 Selecting Samples 25 245 2keeendeidesasetaneeesGerancsehen aes aa 84 Creating a Custom Report Template 20 0 cece eee eee 88 e MA eee seen ede deoes 4 eeee ee eee a r bes 90 Specifying Report Paper Bi ZG 24 94 44 4409 94 495949 aca 92 Chapter 9 HLATools Legacy Reporter Preformatted Reports sports cess bene eles eee eaten Dri 93 Patient Reports 2 seeds wide recrea 94 1B Patient Summary Report Filtered by Batch 94 2A Patient Repo ciosscrserisnrcror adi o 94 Bali Repos 22axegida der sosa candado aro 95 3A Batch Report lt icascccesbaritdcdderauneuberabaradeevakacihs 95 3B Batch Raw Data Abbreviated Specificity Report 96 3D Batch Report Sorted by Patient 0 0 0 e eee eee ee 96 3E Batch Report Sorted by Sample o 2444 2c0 e0de lt idee es targa cede 97 3F Combined Batch Report Sorted by Sample 97 3G Combined Batch Report Sorted by Patient 98 Sampl REPOS td 4 aha A ede at Ede aan 98 3C Sample Report Filtered by Batch o oooooooooooo 98 4A Sam
31. Anthony Nolan website no alleles have been named with the A or C suffixes Version 1 1 Table B 3 HLA Allele Nomenclature Examples Nomenclature Meaning DRB1 DRB1 11 DRB1 1101 DRB1 1160N DRB1 110101 DRB1 110102 DRB1 110103 DRB1 11010101 DRB1 11010102N DRB1 1101 03 DRB1 11MN DRB1 1101 03 DRB1 11AC Locus Group of alleles that encode the DRB1 11 antigen Specific allele Specific null allele Allele that differs by a synonymous polymorphism Allele that differs by a synonymous polymorphism Allele that differs by a synonymous polymorphism Allele containing a variation in a noncoding region intron polymorphism Null allele containing a variation in a noncoding region intron polymorphism Ambiguity DRB1 1101 or DRB1 1102 or DRB1 1103 The above using NMDP ambiguity code Ambiguity DRB1 1101 or DRB1 1103 The above using NMDP ambiguity code OLI UD 0157 062006 117 of 172 HLATools_IVDManual_v1 1_ Rev 0 pdf Matched Reaction Pairs MRPs How does LTI identify Figure B 4 illustrates how LTI determines Matched Reaction Pairs pairs of alleles whose combined reaction patterns exactly match the observed sample reaction pattern Close Reactions pairs of alleles whose combined reaction patterns differ by one bead reaction positive or negative from matching the observed sample reaction pattern and False Reactions pairs of alleles whose combined reaction patterns differ by one or more bead
32. B 57XX4 may be B 570301 or B 570302 2 possibilities Together these yield 10 pairs as indicated by the cells in Figure B 9 marked with rectangles A complete matching of the possibilities from the cells in both columns yields the twenty pairs contained in the table of Figure B 9 Figure B 9 Recognized Allele Pairs Serological Equivalents Recognized Allele Pairs Serological Equivalent Allele1 Allele2 Filter See ee ras safe Jee or Jor jews 5500 CON CON EN CN CE 5000 CON CONAN CI CAS EX E e a7 ow few ewes eevee e ey ows fot eres Cc oJ foe fo B 570302 GUCCI CA COCA CO on Jao ares Dc E os EX ll E fr om ows CS CA CA A O CO CA CC pares Berar ee font os pybads suxy 3585015 adAqgns ad4l sajaypy jy PAYNEN 124 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Type Subtype Allele Assignments How are made This FAQ addresses how the Type Subtype table is generated from the entries in the All Alleles table Table B 4 1 All digits beyond the first four are dropped columns 1 and 2 in Table B 5 2 All resulting duplicate pairs are discarded columns 3 and 4 3 All duplicate entries in each column are discarded columns 5 and 6 The resultant columns are identical to those shown in the Type Subtype listing Figure B 10 Table B 5 Creating the Type Subtype Tabulation Truncated to 4 Digits Duplicates Pairs Discarded Duplicate Entries Disc
33. Clustering Positives to the Left of the Grid You can group all the positive reactions for a given allele to the left of the table by clicking in the empty cell directly to the left of the allele designation Figure 5 20 Note the revised bead order in the top row of the table in ascending numerical order 6 7 11 Version 1 1 61 of 172 OLI UD 0157 062006 HLATools_ VDManual_v1 1_Rev 0 pdf Figure 5 20 Clustering Positive Reactions to the Left 1120222324 3681 2 3 4 5 8 9 10 12113 14 15 16 17 18 43 PP Alleles Oe A A ie x x x x x 0 Allele 6 7 Run x x N DPB1 03CXD S DP81 5201 DPB1 1301 DPB1 2901 0 DPB1 030101 0 DPB1 5201 0 DPB1 1301 0 DPB1 2901 0 DPB1 7001 O DPB1 8801 0 DPB1 260102 0 DPB1 1401 0 DPB1 5001 DPB1 7001 DPB1 8801 DPB1 260102 DPB1 1401 _ DPB1 5001 x xX xX KK KK OK OK Restoring Allele Order You can restore the bead order to its original state by clicking in the upper left cell of the table just to the left of the Allele column header Figure 5 21 Figure 5 21 Restoring Bead Sort Order LN Alte et 2 3 4 5 617 8 9 10 19 12 13 14 15 16 17 18 19 20 22 23 24 25 35 36 Rxn x x eae x A x X XXX x DPB1 03CXD DPB1 5201 DPB1 1301 DPB1 2901 DPB1 7001 DPB1 8801 DPB1 260102 DPB1 1401 DPB1 5001 DPB1 5601 x xX xX KK MK KK OK OM Shifting Alleles to the Top of the Grid You can shift an allele ab
34. Connection Name A Configuration Connections mann Add Database Attach Detach Modify Backup Restore Clean Update Merge Diagnostics Delete Test Connection User Management 14 When Quimby next launches LTI from his computer skipjack he will be able to access the LTI database on the target server swordfish via the Homer connection You can access remote databases on any computer on which you have been added to the authorized users list and you can also add and delete users from the remote computer s authorized users list This administrative function is described below in Editing the Authorized Users List from a Remote Server p 146 144 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 15 Quimby can re access his local LTI database by launching the Connections tool and selecting File gt Connection gt Quick Connections gt Local to restore active status to the Local connection or by using the Ctrl L key stroke combination Note that a Ctrl lt gt key stroke combination is assigned to each Quick Connection This allows you to switch between database connections without using the menu In addition to the default Local Quick Con nection Ctrl L you can create up to nine additional Quick Connections Ctrl F1 through Ctrl F9 Figure C 10 Reactivating the Local Serves Connection HLA Tools Interactive Configuration Suite Quick Connections gt EIA Custom
35. Connection w Homer Ctri F1 Po Database Attach Detach Backup Restore Clean Update Merge Diagnostics User Management Modify _teaty Add OK Cancel Apply 16 To summarize the controls the File menu options and the right click menu options in the Connections dialog have the following functions Add creates a new connection Figure C 6 Apply saves the settings without closing the dialog box this allows you to test the connection without leaving the dialog box enabled only when a connection has been selected Custom Connection creates a connection for one time use without saving the connection Delete right click menu option deletes the selected connection Make Active right click menu option activates the selected connection Modify accesses the Database Settings dialog Figure C 7 where you can specify the type of authentication employed by the connection SQL or Windows and define a new connection Quick Connections accesses the local database and up to nine other predefined connections OK saves the settings and closes the dialog box Version 1 1 145 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Test Connection provides confirmation that database settings are valid Editing the Authorized Users List from a Remote Server Once you have established a connection between your computer and a second server you can also use the User Management
36. First Allele Entering a Second Allele in the other sort field moves that allele directly beneath the sample reaction pattern and forces the remaining alleles to be reordered according to their similarity to the Second Allele In this sorting the alleles are Version 1 1 63 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf sorted downward in order of decreasing similarity For a more detailed discussion of the sorting methodology see Reaction Grid How is the sorted p 126 The swap button lt gt between the two sort fields switches the entries See 5 to Figure 5 16 Figure 5 24 Two Way Sort by Allele Pattern Match Allele B 1506 12 B 1506 B 1539 12 B 1539 B 1588 13 B 1588 B 1555 13 B 1555 B 1531 13 B 1531 B 1515 13 B 1515 B 1502 14 B 1502 B 0723 B 074MY N B 0724 B 0733 B D737 B 070202 B 0710 B 0717 B 0716 B 0720 15 B 0723 15 B 070201 14 B 0724 14 B 0733 14 B 0737 14 B 070202 14 B 0710 14 B 0717 14 B 0716 14 B 0720 x x x x x x x x x om x x x KX KK KM KK KK KK OK ox Re SS eM Meee Ne oe Me O ee O O K KKKA x xX KX KK KK RK ox x xX KX KK KK KK MK OK x xX KK KX KK KK OK OK x xX KX KX KX KX KK KK OK x KX KX KK MK KK KK OK x x xX KKK MK OM x xX KK KK KK KK OK x xX KX KK KK KK MK ox x xX KK KK KK KK OK This bidirectional pattern sorting is a powerful tool for clustering allele reaction patterns An allele with a high density of positives at the front end o
37. Modules ida ib A RA AA A 7 HLATools Configuration Suite 0 0 ccc eee ee 8 HLA Updat AAA 8 Comments Editor iii da as 8 CONMSUTANOM AAA AAA AAA ARA 9 Connections 60 242 0 rers tasa ses aaee rise genes 9 Database 16018 receure rede nren iati rrie rade n hee eana R eane a 9 System DIGGNOSUCS so nt cae eee d eae ee psa eee ho eagy aoe ok 9 User Management 36 sn 34 6 454 AA OGG Ek bee Rae bee Eade 10 E MAA 10 About this Manual dirt AAA 11 How to Use This Guide crop AAA AAA 11 Viewing the HTML Version of this Manual 0 00000 5 12 Conventions Used in this Manual 20 0 0c eee eee ee 12 Inmt rtace NomenCl Aare sss den as Sw dd ES SASS RONDA a 12 Technical Support ii A A A 13 Chapter 2 Installation and Log On Software Installato ere ede t eee nee ee Sek a TEn EREET ee Cees EEEE E REGE 15 System REQUINGMICIISS 4 006 249 645049444 49046955 II 15 Luminex Software Compatibility 0 0 0 0 0 cece eee eee eee 16 Software Installation on all User Computers 0 0 0 0 0 0 ee eee 16 Creating the LuminexFiles Folder 2 205 2 60e048 4dsree8 04 as 18 Running the Clean Database Utility cocer AAA 18 Modifying Repairing or Uninstalling HLATools oooocoococococooo oo 19 Modifying HLATOOIS 2 020 ee ee eee dee ee sede eee eea dese rra 20 Repairing HEA TOONS 4 dada adds dan 21 Version 1 1 HLATools User Manual 1 of 172 OLI UD 0127 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Chap
38. Positive Negative False Reactions posited by LTI when making suggested allele assignments in Close Reactions 15 Y axis Bead ID numbers in the batch double clicking on a bead ID number opens the Bead Analysis view for that bead 16 Tool tip dwelling over a histogram column displays the bead ID and the count of Close Reactions in which the bead is involved double clicking on the column opens the Bead Analysis view for that bead Note that Mousing over a histogram column for one type of false reaction causes the columns for the opposite type of reaction to become dim This makes it possible to determine when false positives and false negatives for the same bead are superimposed one upon the other 17 Locked when checked prevents the bead from being excluded from analysis and any modifications of the cutoff value 18 Print prints the histogram currently displayed in the main panel QC Data or Sample Data Version 1 1 43 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Control Values View Positive Controls The positive control values are used in the normalization calculations for the individual bead fluorescence values The control values are actual fluorescence readings The formula used to normalize the bead readings can be found in the glossary 1 Positive Control Values the Y axis indicates the fluorescence value of the positive controls There values typi cally range from 1000 to 4000 LTI autom
39. SQL Server network protocols enabled If not the program will enable it This occurs without user intervention 5 The installation program next checks to see if an SQL database exists on the server instance If an SQL database does exist the program makes a backup copy If one does not exist the program will create a new database This may require several minutes 6 At this point a window similar to that shown in Figure B 16 appears which lists the file paths for the LABType ses sion files Click Upgrade to continue Figure B 16 Upgrade LABType Database Files Upgrade LABType Database iol xj ClMy Sess DO Project lxp CALUMINEXFILES CSY FILESIDRB1_LOTSTEST1 CSY Upgrade CALUMINEXFILESYOUTPUT CSY K PRODUCT SPECIALISTILABMAS WORKSHOP JANUARY ZOOHDATAIWSO4BLOC K PRODUCT SPECIALISTILABMAS WORKSHOP JANUARY 2004 DATA1WS04ALOC CHULUMINEXFILES1041204BL62 C5W CHALUMINEXFILES1041204BL62T C5Y Version 1 1 133 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 7 The utility performs the following steps for the samples in each batch A Imports raw data count and trimmed mean B Normalizes the data C Copies the LABType analysis results into the SQL database and maintains the Locked status of samples whose allele assignments have been reviewed and accepted D Carries over any user adjustments in bead cutoff values and notes that the cutoffs have been changed from the default values E Carries
40. See Gh x 5 0 1 1 12 A 0303N 2 Sie MO E SS PS X 5 0 1 1 11 A 0306 Nat oe Xa Neg Ke KD eS 5 0 1 1 11 A 0308 x SGC GT A R ep RE K 5 0 tl 2 11 A 0309 A SG Os ee eek ee O de a 5 0 11 2 11 A 0312 KEKER te OCR RIR Ce 96 AAK 5 0 11 2 11 A 030102 A gece Saw Cw OS a 3 0 11 1 11 A 03ANGK x x z xax Ke x Xe A 3 0 2 1 2 pan AE E AX EX A 68NPN re BE Oe SE Be 5 E 0 5 7 1 12 A 6815 x ak X X 0 0 7 3 10 A 6827 2G Be RS FS We x UE 0 0 6 2 11 A 3403 ee eee SE ES x Seay X 0 0 0 2 10 A 6823 eye As aes 0 0 0 2 10 NMDP Codes How can automatically update The easiest way to assure that HLATools is using the most current NMDP codes is to update them automatically Windows XP and Windows 2000 both include a utility that you can use to set up an automated task or chron job to update the codes In an ordinary installation of HLATools the NMDP updating utility InputNumericTxt exe is located in C Program Files One Lambda HLATools You can use the Windows Scheduled Tasks utility to run this executable automati cally The following steps describe the setup in Windows XP Some terms may be slightly different in Windows 2000 1 From the Windows desktop select Start gt Settings gt Control Panel gt Scheduled Tasks 2 Launch the Add Scheduled Tasks application 3 The Scheduled Task Wizard opening dialog will appear Click Next to proceed 4 Locate and select Update NMDP Data in the applications listing or you can browse to locate the ut
41. Selection panel In the Search panel select the Sections tab then Edit The sections included by default in the existing Sample Report are checked and in bold face type Figure 8 11 An actual report containing these default sections is shown in Figure 8 13 e When you have included or excluded the desired sections in the template Save the template to the folder con taining the templates of that report type You will be prompted to rename your custom template so you do not write over the stock template Your new report template is added to the listing of reports of that type Figure 8 12 e Creating templates for other types of reports differs only in minor details 88 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 8 12 Batch Reports Listing Batch Reports Batch Report MyCustomB atch Report Figure 8 13 Default Sample Report Default O 5102006 Page 1 of 1 YourLab Your Street Your Town CA 90000 y Your Director Tel 800 555 1212 Fax 818 555 1213 E mail you yourlab com _ www yourlab com LABType SSO Class II Data E Sample ID Sample 5 Catalog Lot RSSO2B1_ 008 Rev 03 Locus DRB1 Batch Name ug04WorkshopDRBILocusTest Analysis Date 5 3 2006 Tested By Test Date 712912004 File Path C LuminexFilegDRB1Lot008 csv Batch Analysis Comment Patient ID Szislack4231 Gender Mi ABO Rh Patent Name Szislack Moe Race C Comments Corrected Typing DRB1 01BREY DRI DRB1 14
42. access to the C MyBatches Output folder map this folder as a drive from the HLA Tools PC 1 Before you start determine the network identification of the Luminex 100 PC e On the Luminex 100 PC select My Computer gt Properties gt Computer Name This latter tab is called Network Identification on Windows 2000 e Note the Full Computer Name and the Domain 2 On the HLA Tools PC e Select My Computer gt Map Network Drive From the Drive pull down menu select an unused drive letter e Select Browse gt My Networks Places gt Entire Network gt Microsoft Windows Network gt lt network branch gt If you do not know the name of the network branch where the Luminex 100 PC is located ask your system administrator e Open the node with name of the Luminex 100 PC select the Output folder and click OK to map the drive The Output folder will now appear in the listing of Network Drives when you open My Computer on the HLA Tools PC Figure C 19 Version 1 1 153 of 172 OLI UD 0157 062006 HLATools_ VDManual_v1 1_Rev 0 pdf Figure C 19 Mapping C MyBatches Output on a Remote Computer Browse For Folder zi xd Select a shared network folder My Network Places E Entire Network E a Microsoft Windows Network E gy YourLab E Y 124 MiRed 4 125 MiDrange 4 129 MrLumined 2 Output E Printers and Faxes Y Make New Folder OK Cancel You will now be able to browse
43. accessed via the HLATools Configuration Suite For an explanation of allele nomen clature see the FAQ topic Alleles How are they designated Test reactions are expressed as a percent of the normalized raw reading according to the formula Raw NC Data 10 PC_NC GI 1 where Data normalized fluorescence reading shown in most tables Raw raw fluorescence reading value PC positive control value NC negative control value 162 of 172 OLI UD 0157 062006 HLATools User Manual 6 6 06 HLATools_IVDManual_v1 1_Rev 0 pdf Nucleotide Codes Null Allele PBS PCR SSP Version 1 1 OLI UD 0157 062006 The following nucleotide codes are recognized by the International Union of Biochemistry IUB and the International Union of Physical and Applied Chemistry IUPAC Table Gl 4 IUB IUPAC Nucleotide Codes IUB IUPAC Meaning Description A A Adenine B CorGorT Not Adenine Cc C Cytosine D AorGorT Not Cytosine G G Guanine H AorCorT Not Guanine K GorT Keto M Aorc Amino N X Aor C or GorT Unknown R AorG Purine S CorG Strong T U T U Thymine Uracil V AorCorG Not Thymine Uracil Ww AorT Weak Y CorT Pyrimidine Comparative studies between serology and molecular typing for HLA A and B loci have discovered alleles detected by DNA typing but not detected serologically These HLA expression variants commonly referred to a null
44. and the user s IT department HLATools is not designed to be an automated network configuration tool For wireless network installation the user must ensure electromagnetic compatibility and is responsible for electromagnetic interference Peer to Peer Installation In order to use LTT in a peer to peer or network configuration it is necessary to enable two communications protocols on each computer Named Pipes a protocol for exchanging information between two applications possibly on different computers and TCP IP Enabling these protocols requires administrative user privileges To enable Named Pipes and TCP IP 1 From the Windows XP or Windows 2000 desktop select Start gt Run 2 Enter svrneten in the Open field and click OK to access the SQL Server Network Utility Version 1 1 135 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Note If you have applications on your system that use versions of the SQL Server Network Utility earlier than 8 0 executing the Run gt svrnetcn command may launch the older utility If this occurs navigate to the newer executable which can be found in C Program Files Microsoft SQL Server 80 Tools Binn and run svrnetcn exe from that location Figure C 1 SQL Server Network Utility SQL Server Network Utility i x General Network Libraries Instance s on this server swordfish ONELAMBDA Disabled protocols Enabled protocols NWLink IFSP Named Pipes Enable
45. antigens with shared Epitopes 158 of 172 HLATools User Manual 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf EDTA ELISA Epitope False Negative False Positive False Reaction Version 1 1 OLI UD 0157 062006 Ethylene Diamine Tetraacetic Acid a chelating agent used as a buffer in preparing blood samples Enzyme Linked ImmunoSorbent Assay the antibody detection method used by the Lambda Antigen Tray LAT assay The LAT assay is based on the principle of presenting purified HLA antigens as targets for the binding of a patient s antibody In LAT defined amounts of purified HLA antigens are presented in the different wells of a Terisaki tray The specific binding of serum antibody with any of these antigens is detected by the subsequent incubation with alkaline phosphatase conjugated antibody that recognizes only human IgG A quantitative measure of the extent of reaction is obtained by spectro photometric determination following the addition of an enzyme Substrate appropriate for the development of color Qualitative assessment of antibody specificity is performed by the analysis of the LAT reactivity pattern A unique shape or marker on an antigen s surface that triggers a corresponding antibody response Epitopes are classified as private or public e Private epitope an individual HLA specificity e Public epitope shapes or markers that many antigens have in common as a result of havi
46. ap parent need for a conventional username password combination should dissuade passersby and the idly curious from accessing the application Users should be aware that usernames are included in analysis reports The security mode employed on a given computer can be set using the HLATools Configuration utility To set or change a security mode setting Select Start gt Programs gt One Lambda gt HLATools Configuration Suite gt Configuration to access the HLATools authentication options Figure C 15 Version 1 1 149 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure C 15 HLATools Configuration CAS Tools Interactive Configuration Suite f oj xj File Analysis Comments II Luminex Files Folder Configuration c LuminexFiles 3 E Connections Database Reports Folder Attach Detach C Program Files One Lambda HLATools Reports Browse Backup Restore Clean Update LabScreen Folder Merge c LABScrn ue Diagnostics User Management Export Type Default Character Encoding Type Janst bi Login Type None Windows Local OK Cancel Apply Any change made to the settings will be in effect the next time you launch LTI Sharing Luminex Files on a Network In a typical laboratory workflow Luminex 100 output files are stored on a computer attached directly to a Luminex 100 Analyzer The output files are often transferred
47. can represent more than 65 000 unique characters This utility rebuilds the LTI database with new product information when the user updates the HLATools software In contrast to Clean Database which creates an entirely new database Update Database maintains all the analysis results and user input that was contained in the old database HLATools User Manual 165 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf This page intentionally left blank Index Symbols slash alternative allelic subtypes 73 tilde range of allelic subtypes 73 self evident filepath name 91 A A 02 A 92 allele family 155 Al 4Digit A2 4Digit 38 A260 A280 Ratio 155 ABDR locus reports 6 Accept function re enabling 47 requiring review by two analysts 47 108 ACD acid citrate dextrose 155 Active content disabling browser warning message 10 Allele name extensions 155 Ambiguity codes local 120 160 NMDP 120 Amino acid codes 155 AO acridine orange 157 Application Data hidden folder 20 ASO allele specific oligonucleotide 156 Authentication mode SQL 139 147 Windows 139 Authorized users list editing 146 B B 15 B 95 allele family 155 Batch analysis Version 1 1 OLI UD 0157 062006 forcing LTT to reanalyze batch 35 BCIP 165 Bead Analysis Histograms viewing entire width 109 Bead Probe Information data sheet 73 Bead specificities copying to clipboard 54 displaying 54 Bw4 Bw6 sero
48. comments and filter status eXport exports a summary text file containing Sample ID Bw serotype equivalents and the Sample Reaction Pattern for each sample in the batch BatchResultsGrid each row contains information about a single sample Tool tip contains abbreviated text of the sample comments such as e False reactions beads whose cutoffs would have to be changed are shown on the left the resultant allele as signments on the right e Ambiguous calls the type of ambiguity and the alternative calls for each allele Low Bead Count noted if the count for any bead falls below the minimum set in the HLATools Product In formation table Figure 7 3 Table Layout Options Menu abbreviated A full listing of the columns that can be included in the Batch Results Grid is accessible by right clicking anywhere in the table itself Information that is always included by default is indicated by an asterisk in the Columns comments below Customer specific information is indicated by a double dagger Customer specific display of results is addressed in Specifying the Data Transfer File Export Type p 108 Column order can be modified as discussed in Appendix A LTT Interface Customization Right click menu options e Save saves the selected columns and column order Version 1 1 37 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf e Load reloads the previous columns and column order thus undo
49. dialog 148 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Setting up HLATools in a Client Server Network To install HLATools in a client server configuration where Windows Enterprise Network with Domain Based Security is being used should follow these steps Log on with Administrative access Install HLATools Software Reboot the computer once Log on again with Administrative access Provide SQL access privileges to user Log off as administrator Log on as User SoS Oy E m Continue with the actual installation of the software HLATools Security Modes Three security modes have been implemented to control access to HLATools In all cases a record of login attempts can be kept with username time and date of login being saved to a log file accessible only by users with administrative privileges In all three modes access to the local computer on which the HLATools suite is installed requires a normal Windows login to boot up the computer None any user can access HLATools without further login e Windows a user must log in to HLATools using his or her standard Windows username password combination This prevents one user from running HLATools under another user s account e Local any user can access HLATools by entering the same string for both username and password in the HLATools login dialog The laboratory management can inform users as to the username password format The
50. in analysis parameters that may affect allele assignments can be made only when the sample is unlocked For more on invoking customer specific functions such as double acceptance see Specifying the Data Transfer File Export Type p 108 16 Patient Information patient specific information is entered in the Patient Information subview The same patient information can be associated with multiple samples in one or more batches 17 Patient tab to access the Patient Information subview 18 Pos Ctrl histogram of the raw fluorescence values for the positive control beads Class I products type alleles in two exons so they have two positive controls Class II products type only one exon 19 Arrow collapses expands the Positive Controls histogram collapsing the Positive Controls panel provides more screen space to view the Reaction Pattern Histogram 20 Reaction Pattern Histogram displays normalized bead readings for the sample reaction pattern Allele assign ments and alerts to conditions such as low positive control values or low bead counts are displayed in the header field Selecting Allele 1 or Allele 2 or both in the MRP table displays the corresponding assignment s The Reac tion Pattern Histogram allows the reviewer to adjust bead cutoff values on a local basis and view the resultant changes in suggested allele assignment It is particularly useful for visualizing what adjustments would have to be Version 1 1 49
51. information only for the specified sample in a single specified batch this report contains the information for all samples with the specified sample ID in all active batches 4D Sample Abbreviated Specificity Report This report contains information identical to 4C with the exception that the tabulated raw data contain only the following three entries e Bead number e Reaction status 1 or 8 e Abbreviated allele specificities for each bead 4E Sample Complete Specificity Report This report contains information identical to 4C with the exception that the tabulated raw data contain the complete i e non abbreviated allele specificities 99A Sample Report with Chart Filtered by Batch This report contains the following information for each sample in specified batch plus a Reaction Pattern Histogram e No patient information e Batch product lot information and locus type for the batch e Draw test and analysis dates for each sample e Session ID and csv filepath for each batch e Batch comments e Corrected accepted typing if any for each sample e No reaction assignment NIH 8 1 format e Computer assignment combining all MRP designations into a single NMDP code pair if applicable e Generic allele descriptions with complete listing of types and subtypes for each MRP e Listing of close reactions e Expansion of NMDP codes for computer allele assignments e Normalized Bar Graph Reaction Pattern Histogram for each
52. is empty LTI inspects the sample reaction pattern and possi ble allele assignments When the software determines that the observed reaction pattern would match that of a rec ognized allele if the reaction state of one or more bead s were changed from negative to positive or positive to negative that bead is suggested as showing a false reaction Candidate false reactions typically involve beads with readings close to the cutoff a bit too low or too high so flipping the reaction state is not equivalent to ignor ing or rejecting the observed data Gray regular face sample has been reviewed and accepted Figure 5 2 LTI Batch Sample File Tree E Inbox Aug04 WorkshopCLocusT est ug04WorkshopDRB1LocusTest 123 104B_L6 Augl4 WorkshopBLocusT est Augl4 Workshop4LocusT est Sample 1 Sample 2 Sample 3 Sample 4 Sample 5 Sample 6 Sample 7 Sample 8 H Deleted H Archive 36 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf The preliminary batch analysis results are presented in three different tabbed views Batch Results View Figure 5 3 Control Values View Positive Controls Figure 5 5 and Bead Analysis View Figure 5 4 Batch Results View The Batch Results View provides a tabulation of preliminary analysis results You can jump directly from the table to the Assignments View for a given sample by double clicking on the s
53. makes it appear in the main display 5 Sample Data corresponding profiles for each bead in the assay As the number of samples in the batch increases the Sample Data bead profile should more closely resemble that of the general population If the bead profiles in the Sample Data vary greatly from the QC data it may indicate that the samples in the batch are not representative of the general population or that some sort of systematic error occurred while preparing the samples or collecting the sample data The Cutoff Sensitivity delta value is user modifiable and can be changed in the HLATools Product Information table Figure 7 3 The default delta value is 10 Thus if the Cutoff Sensitivity were reduced to 5 a reading above 105 would count as very positive and a reading below 95 would be very negative 6 Header displays the name of the QC Data file or the Sample Data bead profile whichever is selected 42 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 7 Y Axis cutoff for the bead as a percentage of the bead s positive control value The Y axis is scaled vertically to accommodate the range of the data 8 X Axis sample IDs for the individuals making up the cross section of the reference population when QC Data is displayed or sample IDs when the batch Sample Data is displayed 9 Cutoff default value for the bead cutoff This value is applied globally to all samples across the batch
54. of HLATools software in a peer to peer configuration or client server network environment is discussed in Appendix C Network Installation of HLATools Software Installation System Requirements These are system requirements for installation and use of HLATools software e Microsoft Windows 2000 SP4 or XP SP2 or later operating system e Microsoft NET Framework v 1 1 e Microsoft SQL Server Desktop Engine 2000 SP3a this must also be installed on client computers in a cli ent server configuration e Pentium III with Windows 2000 or Pentium 4 with Windows XP e 256 MB of RAM e 5 GB of available hard disk space e CD ROM drive All software needed to install HLATools on standalone or client computers is contained on the CD ROM in your software shipment The installation of the SQL database server software should be handled by your IT system adminis trator The MS Windows SQL Server software must include Microsoft SQL Server 2000 Service Pack 3a which is available from Microsoft at http www microsoft com downloads Version 1 1 15 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Luminex Software Compatibility HLATools software has been developed and tested to be compatible with Luminex IS 2 1 and IS 2 2 output files produced by the LABScan 100 flow analyzer Software Installation on all User Computers 1 You must have administrator status on the PC in order to install HLATools and the HLAToo
55. of the row clusters all of the Rxn positives to the left Golden Alleles alleles whose known reaction patterns are a subset of the observed sample reaction pattern Golden alleles exhibit only the positives observed in the sample reaction pattern They are highlighted in the table by a yellow background When the reaction patterns of two golden alleles can be added to yield the observed sample reaction pattern the junction is called a Matched Reaction Pair MRP In Figure 5 16 the reaction patterns of DRB1 11APWP and DRB1 1404 together form an MRP Similarly the junction of DRB1 11UC and DRB1 1404 yields a second MRP Pairs of golden alleles yield MRPs only when their combined reaction patterns completely match the positives in the sample reaction pattern The combined reaction patterns of the golden alleles DRB1 11UC and DRB1 1439 do not suffice to form an MRP since neither shows specificity on beads 68 or 51 False Positive Allele DRB1 1404 is an example of an allele that could be part of an MRP if one of the beads in the sample reaction pattern were a false positive in this case bead 10 Figure 5 17 shows another example of a False Positive The allele DRB1 1105 also exhibits no reaction at bead 10 If the observed reaction of bead 10 in the sample reaction pattern were a false positive then the junction of DRB1 1105 and DRB1 1404 would match the revised sample reaction pattern now minus the false positive at bead 10 Together they
56. over sample specific comments F Checks to see if the Patient ID associated with the sample being imported already exists in the SQL database If the Patient ID exists only information from non blank fields is imported i e no existing data is over written by null values Imported patient demographic data includes Patient ID Last Name Middle Name First Name Race Family Name DOB Relationship ABO Rh If the Patient ID does not exist the program uses the Sample ID as a substitute Patient ID and imports the patient demographic data for the sample into the SQL database The steps outlined in 7 are repeated for each session file shown in Figure B 16 134 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Appendixc Network Installation of HLATools This chapter addresses two configurations for installing HLATools software in a networked environment e Peer to peer configuration where several workstations are connected in such a way that one user can access the files and database s of other users e Network configuration where a number of client workstations are connected to a central server The server handles many of the computational and databases functions for the client machines The standard installation of HLATools on a standalone workstation is discussed above in Chapter 2 Installation and Log On Important Note Peer to peer and network configuration is wholly the responsibility of the user
57. plus allele b is the same as allele b plus allele a the combining comparing is only done for the first case As a result the total number of combining comparing steps is n x n 2 where n is the number of alleles recognized by the LABType product The combining comparing procedure is executed when the user accesses the Assignments View for a given sample either by clicking on the sample in the Batch Sample file tree Figure 5 2 or on the sample row in the Batch Results Table Figure 5 3 2 Close Reactions LTI identifies Close Reactions by toggling the bead reactions in the Sample Reaction Pattern one by one from positive to negative or negative to positive For each change in bead state LTI repeats the com bining comparing procedure described above looking for allele pairs whose combined reaction patterns match that of the now modified Sample Reaction Pattern The search for Close Reactions is executed when the user accesses the Close Reactions subpanel Figure 5 13 in the main Assignments View or the Bead Analysis View Figure 5 9 For each Close Reaction detected LTI reports the allele pair and ID of the involved bead in the Closest Reactions subpanel Figure 5 13 3 False Reactions LTI searches for False Reactions by looking at all allele pairs with combined reaction patterns that are one bead reaction removed from yielding a Matched Reaction Pair or a one away A one away False Reaction is also a Close Reaction
58. reaction positive or negative from matching the observed sample reaction pattern Before starting any of these procedures LTI compares the normalized mean fluorescence values for each of the beads in the sample to the established cutoff values for the beads to determine which beads have registered positive reactions Based on this comparison LTI constructs the Sample Reaction Pattern for the current sample see Figure 5 8 Assign ments View with two Matched Reaction Pairs The reaction patterns for all samples in a batch are determined when the initial batch import in done 1 Matched Reaction Pairs LTI makes an exhaustive comparison of the combined reaction patterns of all pairs of alleles recognized by the selected LABType product to the observed Sample Reaction Pattern The two alleles in each comparison are designated Allele I and Allele II e LTI steps through the entire list of alleles combining the reaction pattern of the first Allele I with each of the other alleles each in turn considered as Allele II and comparing the combined reaction patterns to the Sample Reaction Pattern SRP If any combined pair matches the SRP that pair is recorded as a Matched Reaction Pattern MRP This may be thought of as an inner comparison loop e LTI then treats the next allele as Allele I and repeats the combining comparing procedure with all of the other alleles This may be thought of as an outer comparison loop Since any given allele a
59. redispersed The specificity of the coupled monoclonal antibody determines the type of cell collected The One Lambda FluoroBeads products employ immunomagnetic beads Demographic information on allelic representations in ethnic Japanese In LTL the Japan Rank A is the most restrictive while Rank ABC is the least When no NMDP ambiguity code is available to designate a group of allelic subtypes with a common reaction pattern LTI designates the group with a temporary local code usually of the form XXn As soon as an appropriate NMDP code has been defined and the internal LTT table of NMDP codes as been updated the local code is replaced by the NMDP code Log files hold all of the log information used to recover the database There must be at least one log file for each database although there can be more than one The recom mended file name extension for log files is Idf See also Primary Data Files mdf 160 of 172 OLI UD 0157 062006 HLATools User Manual 6 6 06 HLATools_IVDManual_v1 1_Rev 0 pdf Matched Reaction Pair Mean and Median Merge Database MIC N80 K80 Negative Control Version 1 1 OLI UD 0157 062006 A pair of alleles whose joined bead reaction patterns match that of the observed sample reaction pattern If the patient is heterozygous for the sequence being typed each of the two alleles will come from a different parent If the patient is homozygous the alleles from each parent will be identica
60. replaces any edited values Cutoff Sensitivity establishes the sensitivity delta percent value that distinguishes near positive or negative bead readings from stronger positive or negative values The default Cutoff Sensitivity is 10 The distinctions between the different reading ranges are illustrated in Table 5 1 Bead Analysis Color Codes Save saves edited values to the database overwriting previous values Cancel undoes any unsaved edit made to the table Figure 7 3 HLATools Product Information ProductName QABatchID Loci Lot Revision R5502345 15 DRB2345 002 R2 RSSO2B1 A DRB1_ DRB n L RSSO2B1 ie DREI a CountType MinBeadCount MinPCData ASSO2B1 EN EA DRET l Trimmed Mean Count D RS502B1 DRB1 Tiersen IS EC LI CI R5SO2QB1 DQB1 Finnedean count a a ACI TI roo 0000 CI Frinnedean cout i00 a The columns in the Product Information table display the following information Any column in the table can be sorted alphanumerically by clicking on the column header The sort order can be reversed by clicking a second time Column subsorting can be invoked by shift clicking on additional column headers For example clicking first on the ProductName column header sorts all products by catalog ID Shift clicking on the Lot column header then subsort all products with identical catalog IDs by ascending lot number ProductName the One Lambda assay catalog ID The product name typ
61. that you include in the sample specific comments is time stamped and shows your username You can edit these stock phrases using the Comments Editor utility To edit the stock phrases Version 1 1 107 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf e Select Start gt Programs gt One Lambda gt HLATools Configuration Suite gt Comments Enter a new comment in the Comments field in the first line of the listing or edit one of the existing comments and OK your edits The changes will be available when you restart HLATools Figure A 3 HLATools Comments Editor Tool E HLATools Configuration Suite Of x File HLA Update User Comments Configuration Comments LabType Labscaen Retest Required Connections Save two copies of the report For this sample System Diagnostics User Management OK Cancel Apply e You can delete a comment by either of the following means e Highlight the table row by clicking in the left most column press Delete from the keyboard and follow the prompts or e Select the comment text press Delete from the keyboard then Apply the row will disappear from the User Comments table Specifying the Data Transfer File Export Type Some institutions may wish to employ proprietary data transfer formats when exporting results from HLATools Unless otherwise specified the default export file type is compatible with
62. the Lambda Report Paper Size tab Figure 8 2 Version 1 1 91 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf This page intentionally left blank Chapter 9 HLATools Legacy Re p O rte r Legacy Reporter The HLATools Lambda Reporter provides tools for generating five categories of preformatted reports in either U S standard 8 x 11 or A4 210 x 297 mm sizes Templates for the preformatted reports in both sizes reside in the C Program Files One Lambda HLATools Reports or ReportsA4 folders The logo which appears in prefor matted reports is located in the file C company_logo bmp and should conform to the logo dimensions cited in Laboratory Report Header Information p 82 HLATools automatically selects the appropriate paper size according to the Regional and Language Options settings in the Windows Control Panel If you wish to use the non default paper size you can override the automatic setting as described in Specifying Report Paper Size p 91 Preformatted Reports The Lambda Reporter generates five categories of preformatted reports which are available in US Standard and A4 paper sizes as discussed above To use a preformatted report 1 Launch the Lambda Reporter By default the Lambda Reporter displays the Filters view 2 Select Report Category gt Report Type e g Batch Reports gt Batch Report Sorted by Sample 3 Invoke the appropriate filter s from the pull down lists as shown Figur
63. the LTI database Export CSV exports table in Comma Separated Value format that can be reimported into the LTI database Serological gt Update Updates the Serological Information table by accessing a serological equivalents file stored locally or on an internet website Filter gt View The Filter View and Update features are currently under development Version 1 1 79 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf This page intentionally left blank Chapter 8 HLATools Lambda Reporter Ure Ponce The HLATools Lambda Reporter provides tools for generating customized reports Report formatting options include e Customization of header and footer information e Inclusion of your laboratory s logo in the header in two different size formats e Five categories of predefined reports or report templates You can edit many of these predefined templates and save the customized templates that you have created The report template categories are e Patient Reports e Batch Reports e Sample Reports e Combined Sample Report e Sample Report e Summary Reports e Batch Results Report e Catalog Information e Data File Summary Report e Database Information Report NMDP Allele Code Report e Patient Typing Summary Report e Serological Equivalent Report e Special Reports Version 1 1 81 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf ABDR by Batch Report ABDR by Batch Report E
64. the sample ID in the Batch Sample tree is displayed in regular non bold typeface Version 1 1 53 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf 4 Previous Next navigation tools for moving to the All Alleles tables of adjacent samples 5 Patient demographic data this information can be edited in the Patient subview accessible via the Patient tab 6 Patient accesses the Patient Information subview 7 Filtered types and subtypes applying a demographic filter to the sample outlined in red results in a smaller number of subtypes in Allele 1 and consequently different ambiguity codes in the allele assignments Type Subtype Subview This subview tabulates the allele types and subtypes for each allele The side by side placement of the two lists is not intended to imply any allele pairing The method used to construct the Type Subtype lists is discussed in Type Subtype Allele Assignments How are made p 125 Figure 5 12 Type Subtype Subview DNA B 44AJVH B 4801 O B 44AJVH B 4801 Print Allele Allele 84402 p 8 4801 Filter Serological Restore E a B 48 Bw Bw4 Bwe JP Locked Commens aaa No False Reactions 3 Matched Reaction Pairs B 444JVH amp B 4801 gt B 44020101 B 440201025 B 4419N B 4423N B 4427 B B 4801 2 4433 B 4434 amp B 4801 R 44N2na amp R 4AN11 gt R 44N2NA amp R 4AN11 Ba Allel
65. to a second computer for analysis using HLA Tools Figure C 16 Alternatively the HLA Tools user can read the Luminex output files directly from the Luminex 100 PC In this section we shall discuss how to set the file sharing properties to allow direct access to the Luminex output files The output files are typically stored on the Luminex PC in a folder called C MyBatches Output User groups or individual users can be granted Read permissions for this folder to enable them to read or copy the output files In the Windows environment the root directory is accessible only to administrators The folder C MyBatches Output is one level above the root directory so it can be made accessible to non administrators if we set the permissions properly Windows 2000 and Windows XP allow you to set user permissions on a per folder or a per file basis Setting the permissions for the folder automatically sets the same permissions for all files and subfolders It is also possible to set more restrictive permissions for a contained subfolder or file The following discussion reflects the Windows XP environment Terminology for Windows 2000 may differ slightly To set the permissions on the C MyBatches Output folder on the Luminex 100 PC 150 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 1 Navigate to C MyBatches 2 Right click on the Output folder and select Properties gt Sharing Figure 3 The default setting for mos
66. would form a Matched Reaction Pair Allele Code Definition clicking on any allele displays a full expansion of the allelic subtypes included in the code the subtypes are displayed in the grid header field 3 and also in the right most column of the table 15 False Negative allele Figure 5 18 shows an example of a False Negative allele Allele DRB1 1109 exhibits a positive reaction at bead 82 that is not observed in the sample reaction pattern If the missing reaction of bead 82 in the sample reaction pattern were due to a false negative reading i e a positive reaction should have been observed then the junction of DRB1 1109 and DRB1 1404 would match the sample reaction pattern which would now also display a positive reaction and form a Matched Reaction Pair LTT searches for and suggests candidate alleles for assignment by looking at so called false reactions These may be either false positive and false negative alleles with known reaction patterns not too different from the observed sample reaction pattern as seen above in Figure 5 17 and Figure 5 18 LTI first considers so called Close Reactions allele combinations that are just one false reaction positive or negative removed from the observed sample reaction so that flipping the reaction state of a single bead would yield a Matched Reaction Pair Then it looks at suggested combinations with two false reactions then three and so forth Bead ID the bead ID number prov
67. 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 2 The Clean Database utility takes several minutes to create and populate the database structure Its progress is dis played in a log message window Figure 2 4 Clean DataBase Log Message Window c CleanDB Starting copy Warning Running Clean Database creates a new clean database which means that all previously analyzed samples and patient information overwritten when the new database is created All current patient sample associations and allele assignments will be lost Modifying Repairing or Uninstalling HLATools The HLATools InstallShield utilities provided on the distribution CD allow you to modify repair or uninstall HLATools You must have administrative permissions in order to perform these functions These utilities affect only the HLATools program files and do not alter your Luminex data files or your database To use the program maintenance utilities 1 Insert the HLATools CD and allow it to AutoStart Alternatively locate and launch the setup utility 2 Click Next to proceed The Program Maintenance dialog will appear Figure 2 5 The InstallShield Wizard allows you to modify repair or remove One Lambda HLATools Version 1 1 19 of 172 OLI UD 0157 062006 HLATools_I VDManual_v1 1_ Rev 0 pdf Figure 2 5 Program Maintenance Dialog jig One Lambda HLATools InstallShield Wizard Program Maintenance Modify repair or re
68. 1 objects selected LTI database directory Why is the growing excessively Your LTT database directory may grow to excessive size if you run Update Database frequently without purging your system of backup database files How this happens becomes clear when we review the steps executed by the HLATools Update Database utility 1 To start the Update Database script creates a new folder called C Program Files Microsoft SQL Server MSSQL ONELAMBDA data backupY Y Y YMMDDhhmm and saves copies of your current mdf and ldf database files to this folder These copies are called simply lt dbName gt mdf and lt dbName gt ldf The default lt dbName gt is LTI This backup copy of the mdf database file is detached from your MSDE SQL server instance to assure that all of its data remains secure The mdf file is the primary data file and the Idf file is a log file 2 Next the script renames the current lt dbName gt mdf and lt dbName gt ldf database files calling them temp lt dbName gt Y Y Y YMMDDhhmm mdf These copies reside in the data folder The temp lt dbName gt Y Y Y Y MMDDhhmm mdf is reattached to your MSDE SQL server instance 3 The script runs Clean Database creating a new database called lt dbName gt mdf in the data folder and merges the data in the renamed temp copy with the new lt dbName gt mdf database At this point the combined size of your database structure will be about three times the original size cons
69. 2 of 172 HLATools User Manual Index 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf
70. 29px for the small logo If you use a long logo the text shown in Figure 8 1 shifts accordingly to the right e Browse to the folder where the logo graphic resides Recommended file formats are bmp or jpg 82 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 8 2 Importing a Custom Logo Search Sections Paper Size Logo Width 0 40 in Height 0 30 in C Short logo Entering Laboratory Information To enter your laboratory information select Start gt Programs gt One Lambda gt HLA Tool Configuration Suite gt Configuration gt Lab Info Figure 8 3 Lab Info Interface E HLATools Lab Information Refresh Save LabID i Cancel LabName Your Lab s Name Phonel 800 555 5555 Address Your Lab s Street Phone gig 555 5555 Fax eta 555 5556 or a 818 555 5556 Address3 fC O O website www yourlab com email yournamei yourlab com City Your Lab s City state ca Zip ooo Labblecior Name ClinicalSupervisor Named LabDirector2 GeneralManager 7 HLASupervisor BillingContact faccountsPayable The Lab Info interface has several controls or fields that warrant discussion Version 1 1 83 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf e Refresh restores all values to those currently saved in the database e LabID can be used to identify multiple laboratories or departments within the same institution e Save saves a
71. 4 05 06 07 apxy 0302 0303N 0306 ADXZ 2301 2302 2303 2305 2306 2307N The NMDP Information View controls are identical in function to those in the Product Information View NMDP gt Update This tool fetches the latest revision of the Code List directly from the NMDP Research website or from a locally stored file Figure 7 9 Updating the NMDP Allele Code List HLA Tools NMDP Update x C NMDP Website NMDP Local NMDP File Location Serological gt View Molecular genotyping is capable of detecting allelic differences down to the single nucleotide or single amino acid level However for some types of transplantation procedures especially those using cadaveric organs allelic genotyping is not possible In such cases expediency requires serological tissue matching Many users may find it useful to relate the results of the LTI allele analysis to their serological equivalent This option accesses the Serological Information table where you can view edit and export the contents of the Allele to Serological Equivalent table Figure 7 10 The revision of this table current at the date of release is provided on the 78 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf distribution CD After the first update the revision date of the code listing is displayed above the table If the Serological table has not been updated since the time of installation the Serological Revision Date label will
72. 5 DRB1 0107 DRB1 0108 DRB1 0111 DRB1 01EW 7 DRB1 14 WHG _ DRB1 14WHG DRB1 1441 DRB1 1446 DRAB1 010103 SS DRB1 1409 DRB1 1406 e Alleles or groups of alleles with reaction patterns that are subsets of the sample reaction pattern are termed gold en alleles and are highlighted by a light yellow background dotted outline in Figure B 6 If the intersection of two golden alleles completely matches the sample reaction pattern the pair is termed a Matched Reaction Pair In the sample shown in Figure B 5 and Figure B 6 the intersections DRB1 01EW DRB1 14WHG and DRB1 010103 DRB1 14WHG both yield MRPs Figure B 7 Figure B 7 Matched Reaction Pairs Matched Reaction Pairs Allele Allele2 DAB1 D01Ew DRE1 14WHG DRB1 010103 DRB1714wWHG Version 1 1 121 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf MRP Table How are Allele1 and Allele2 grouped in the When LTI analyzes the observed sample reaction pattern it creates an exhaustive listing of all known alleles that can be grouped into pairs consistent with the observed Sample Reaction Pattern SRP These pairs are the Matched Reaction Pairs MRPs shown in Figure B 8 However before the MRPs can be shown in the arrangement seen in Figure B 8 the alleles making up the MRPs need to be systematically sorted between Allele1 and Allele2 To illustrate the sorting method we will consider a simple case in which all
73. 7 062006 6 6 06 HLATools_IVDManual_v1 1_Rev 0 pdf e Complete specificity Special Reports 5A Allele Group Frequencies This report contains tables and accompanying graphs of the alleles detected and their phenotypic and genotypic frequencies in the HLA A B Cw DQ DP DR loci analyzed using One Lambda products and LabType Interactive 6D Database Table Information This table contains information about database contents and usage The three columns on the first page of the report list e Run dates grouped by week of the batches currently in the database These numbers include both sample and QA batches The QA batches are used by LTI to construct the QA bead profiles This column currently has no header e Batch number of batches run during that week e Sample total number of samples in the batches Inbox Deleted Archived current disposition of the user s sample batches in the LTI database e QA Batch number of QA batches used to construct the QA bead profiles The information tabulated on the second page is of diagnostic interest to the database administrator 99B ABDR by Batch Sorted by Well Position This report is identical to 1A except that it returns results only for Allele1 It also includes serotypic equivalents for the assigned allele Version 1 1 103 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf This page intentionally left blank AppendixA LTI Interface Cu
74. 7 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf 6 OK the changes You should now be able to connect to the remote LTI database Local security policy on XP How do I modify You must have administrative privileges to change the security settings 1 5 6 From the Windows XP desktop select Start gt Settings gt Control Panel gt Administrative Tools gt Local Security Policy In the Security Settings file tree open the Local Policies folder and select Security Options In the Policy Security Setting listing locate the security policy and its setting Network access Sharing and security model for local accounts Guest only local users authenticate as Guest Double click on the name of policy to open the like named dialog box Select the Classic option as shown in Figure B 2 Figure B 2 Changing the Local Security Policy in Windows XP Local Secunty Setting ey Network access Sharing and security model for local accounts Guest only local users authenticate as Guest OK the change Other users will now be able to access the local LTI database using their own authentication Do this on all computers that will be sharing LTI database access Clean Database Why do get a BCP error If you are using HLATools on an non English language version of the Windows operation system your MS SQL Server Desktop Engine 2000 MDSE may not have been updated to the necessary Service Pack 3 product level owing to la
75. 72 OLI UD 0157 062006 6 6 06 HLATools_IVDManual_v1 1_ Rev 0 pdf e Corrected accepted typing if any for each sample e Reaction assignment NIH 8 1 format is included in raw data table e Computer assignment combining all MRP designations into a single NMDP code pair if applicable e Generic allele descriptions with complete listing of types and subtypes for each MRP e Listing of close reactions e Expansion of NMDP codes for computer allele assignments 3E Batch Report Sorted by Sample This report contains only sample specific data Draw dates for individual samples are not included e Batch product lot information and locus type e Test and analysis dates for each sample e csv filepath e Batch comments e Corrected accepted typing if any for each sample e Reaction assignment NIH 8 1 format e Computer assignment combining all MRP designations into a single NMDP code pair if applicable e Generic allele descriptions with complete listing of types and subtypes for each MRP e Listing of close reactions e Expansion of NMDP codes for computer allele assignments 3F Combined Batch Report Sorted by Sample This report contains data for each sample in order assembled from each batch in which the samples appear Draw dates for the samples are not included The csv filepath in the report header is the one selected in the Batch ID filter Figure 9 2 The filepath for each batch is also reported e Batch product lot in
76. ATools_IVDManual_v1 1_ Rev 0 pdf 6A Serological Equivalent Table Report This report contains the same information as the Allele to Serological Equivalent table that can be accessed using the HLATools Maintenance tool See Survey of LTI Data Resource Files p 71 and Figure 7 10 6B NMDP Allele Code Data This report contains the NMDP code numeric expansions sorted alphabetically The codes can be accessed using the HLATools Maintenance tool See Survey of LTI Data Resource Files p 71 and Figure 7 8 6C Batch Data File Summary Report This report contains a history of batch operations since HLATools was installed on the local computer The following information is provided for each batch Database batch ID number Batch session ID Serial number of Luminex machine used to analyze the batch Luminex run date and operator One Lambda product lot used Batch status active deleted or archived Batch filepath 6E Catalog Information This report contains product information for the selected LABType product Product Catalog Number Lot Number Applicable Product Insert Revision Number Data Type employed Mean Trimmed Mean etc Count Type employed Count or Trimmed Count Minimum Bead Count setting Minimum Positive Control value For each bead e Bead number e Associated positive control bead e Abbreviated specificity e Recognition site with amino acid mutation e Cutoff value 102 of 172 OLI UD 015
77. Control Panel gt Internet Options gt Advanced 2 Deselect the Security option Allow active content to run in files on My computer You may wish to re enable this option when you have finished with your HLATools session Conventions Used in this Manual This manual assumes that you are familiar with the Windows operating system and with the standard desktop functions that are common to virtually all software programs e Basic file operations opening saving and printing e Editing functions cutting copying and pasting text e Resizing windows and dialog boxes e Simple manipulation of tables It also assumed that you are familiar with the standard Windows file and folder tree structure and that you know how to manipulate folders and the files they contain Menu options and dialog box selections in step by step procedures are indicated in green text with a right angle bracket marking each step A step may involve actions such as making a menu selection accessing a pane through a tab or checking a check box or radio button For example to access the HLATools application select Start gt Programs gt One Lambda gt HLATools gt HLATools Names of Views Subviews Panels Dialog Boxes and Buttons are shown in boldface type except when they form part of a hypertext cross reference or a navigation path to a particular view subview or panel When used as hypertext they appear in blue or green type Conventions Used in this Manual
78. H 117 A112 25 amp 0252 4 300101 A 300102 A 6806 A 300201 4 300202 A 300203 4 3003 A 3004 A 3006 A 3007 4 3008 A 3009 A 3010 4 3011 A 3012 A 3013 A 3014L A 3015 A 6806 13 A 0281 A 2432 75 ES RISO A109 2 12 a 0281 4 2432 A 250101 A 250101 02 04 4 2913 A 2502 4 2503 A 2504 A 3201 02 04 110 A 2913 A 3201 A 3202 A 3204 A 3205 A 3206 A 3207 4 3208 A 3209 A 3210 A 32110 Expanding the product row by clicking on the at the left side of the table displays a subtable Figure 7 4 containing specificity data for each bead Note that all fields in these subtables are editable Bead ID ID number of sample bead PosCtrlBead ID number of the positive control bead whose control value is used to normalize the reading for the sample bead AbbreviatedSpecificity a compact version of the complete list of alleles that is presented in the CompleteSpeci ficity column The abbreviated specificity list is used in many of the output reports The specificities are abbreviat ed using these rules e Allele families the entries for a family are terminated by a semicolon 74 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf e Alternation sign the forward slash symbol designates an alternative B 1542 44 means subtypes B 1542 or B 1544 e Range sign the tilde designates a range of allelic subtypes B 1542 44 means subtypes B 1542 B 1543 and B 1544 W
79. MSSQLSONELAMBDA Data LTI o9 6 Inline 12 replace the hard coded value LTI with the environment variable STABLES 12 osql S SSERVER SLOGINS q EXIT CREATE DATABASE STABLES 7 To create a database named MY_LTI_DB on a remote server with the IP address 199 199 1 200 using the default null server instance lines 1 12 should appear as below Login would employ the SQL authentication protocol with the username password combination myname MyWord echo off SET SERVER 199 199 1 200 SE TABLE MY_LTI_DB REM SE OGIN E SET LOGIN Umyname PMyWord REM SET SERVER 192 168 1 200 OneLambda net start mssqlS onelambda osql S SERVERS SLOGINS q EXIT DROP DATABASE STABLES net stop mssqlSonelambda REM Del ProgramFiles Microsoft SQL Server MSSQLSONE net start mssqlSonelambda osql S SSERVER SLOGINS q EXIT CREATE DATABASE STABLES 00 ugna o Ww u Aa 7j D ES U U D iw w ct w a E H H Pree NRO Connecting to a Remote LTI Database LTI is often installed in a peer to peer configuration where one user wishes to access LTI databases on another s computer A major issue in peer to peer installations is the proper setting of permissions so that only authorized users can access remote LTI databases To allow a remote user to access a target us
80. Microsoft Excel Please contact One Lambda Customer Support to obtain the identifier to enter in the Export Type field Figure A 4 that is unique to your insti tution The export type identifier can be specified in the HLATools LABType Configuration panel Specifying a unique export file type can be used to invoke some or all of the following report or display behaviors e Custom export file format 108 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf e Differing criteria for sample acceptance and review e g requiring that samples be reviewed by more than one analyst e Alternative designations for allele assignments e g computer allele assignments truncated to type and subtype digits only e Rescaling of Reaction Pattern Histogram and Bead Analysis Histograms so the entire width can be viewed without scrolling e Alternative display of analysis results in the Batch Results Grid Figure A 4 HLATools LABType Configuration Panel HLATools Configuration Suite l m x File HLA Update mLabType Configuration Comments Export Type Configuration force aaa LabScan Character Encoding Type Connections Database ansi z System Diagnostics User Management OK Cancel Apply Specifying the Character Encoding Type Encoding is the process of transforming a set of Unicode characters into a sequence of bytes The Unicode standard assigns
81. Next the installation software prepares for the actual installation of the HLATools software package When the preparation steps have been completed you may be requested to reboot your computer If so click Restart and continue After Windows restarts the next step in the installation may last several minutes 16 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 8 When the HLATools Setup InstallShield Wizard appears click Next read and accept the license agreement and click Next again to continue Figure 2 3 Customer Information Dialog ji One Lambda HLATools InstallShield Wizard Customer Information Please enter your information User Name Your HLATools UserName Organization Your Institution Name Install this application For Anyone who uses this computer all users C Only For me oli Installshield lt Back Cancel 9 Regarding the Customer Information dialog A The User Name field is sometimes filled in automatically by your computer B An entry in the Organization field is optional C Itis recommended that the default option that allows any user of the computer to use the HLATools software be accepted Click Next to proceed 10 Two Setup Type options are available A Complete installs all program features and requires about 260 MB The default installation destination is your C drive B Custom allows users to select installation options
82. On Line Manual HLATools HLATools v 1 1 June 2006 For In Vitro Diagnostic Use N ONE LAMBDA INC 20001 Kittridge Street Canoga Park CA 91303 2801 Advancing Transplant Diagnostics Tel 818 702 0042 Fax 818 702 6904 www onelambda com Manual_Template_R1 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Copyright 2006 One Lambda Inc LABType is a registered trademark of One Lambda Inc HLATools is a trademark of One Lambda Inc LABScan 100 is a trademark of One Lambda Inc Luminex is a registered trademark of Luminex Corporation Windows is a registered trademark of Microsoft Corporation All One Lambda software products are designed to assist personnel experienced in HLA analysis by suggesting typing results However any clinical or diagnostic results must be carefully reviewed by a person qualified in HLA typing to assure correctness The software may be used to aid in suggesting results but should not be used as the sole method for determining reportable results The software is meant as a laboratory aid not as a source of definitive results The software design does not mitigate hazards associated with the software The laboratory director or technologist trained in histocompatibility testing is required to review all data to detect any problems with the software Table of Contents Chapter 1 Introduction to One Lambda HLATools A caceee he baded so E a E E E nace 7 HLA Tals
83. Version 1 1 23 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 2 9 SQL Server and Database Status E HLATools Configuration Suite gt HLA Update Comments E Configuration i Connections E Database i System Diagnostics User Management 2 Ifyou want to be sure that no results are lost you can back up a database using options provided by the Backup Restore utility It is recommended that you use the Backup tool since the database remains attached and available to other users while it is being backed up Even very large databases are typically backed up in less than one minute 24 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 2 10 Backup Database HLATools Configuration Suite File gt 001883 SEB OLD ONELAMBDA HLA Update Comments Configuration Connections Databases Database LTI Attach Detach Local Create Restore From Merge System Diagnostics Path User Management c LTI 2006_6_5 14_48_45 backup cae NOTE When backing up a remote database the path refers to a directory on the remote server not on the local machine Back up To Select the database you want to back up from the pull down list and specify a name and path for the back up file using the browse feature Uninstall the old version of HLATools by selecting Start gt Settings gt Control Pan
84. W es serorcrisdrriaricer terrario 56 Sample Assignment SUBVIEW ceca 56 Patient Information Subview o o ooooococococo eens 57 Bead Analysis Sample os ond ine vee di irse area 58 Reaction Grid AAA inet eed eee eae KE eee eee yee eo uk eles 58 Sorting the Reaction Grid s 4 542cseee Geet sees eendeeaSeereeeesees 61 Clustering Positives to the Left of the Grid oooooooooooooooooo 61 Restoring Allele Order cc 26 ceeds abet iasededeeaerscnreendaues 62 Shifting Alleles to the Top of the Grid 0 0 2 0 eee eee 62 Restoring Allele Order 0c cece eee center tenes 62 Sorting by Bead Usage s c odeaseciderete vadedasetideanne se ies 63 Sorting by Allele Pattern Matches 22 00 eee eee e ee eeeee 63 Raw Data Table 223 625 44 228be hernia sur roer 64 Patient Explorer A A Eadie MNO EEE OEMS Sp eR Ca alee EE eae mee A 69 Version 1 1 OLI UD 0127 062006 HLATools User Manual 2 of 172 HLATools_IVDManual_v1 1_Rev 0 pdf Chapter 7 Reference Files and Their Maintenance Survey of LTI Data Resource Files 004405 4004e0ves eee ete dada a ado 71 Products gt VIEW a A A A A Cabo aba e ds 72 Products gt Update v6 crus be eS es Se ee Gece AAA eee ESS 75 NWIDE gt VIE aye ou ibaa rdd A a a a eae eee 77 NMDPS Updates ccsesergebdeuasd ci deeereteoeea he rider 78 Serolorical VIEW dias Raa stew E See SR eG ea ees 78 Serological gt Update cocina d eee 79 Filter VE aa aa 79 Chapter
85. WHG DR14 Computer Assignment DRB1 01 BREY DRB1 14 WHG Genenc llele Descriptions DRB1 01BREY gt OVO50 ROSI 11213 DRB1 14wWHG gt 02 47 Report Sections The sections available for inclusion in a report are described below The default sections of a Sample Report are shown in Figure 8 13 Other types of reports contain differing collections of default elements Lab Information 1 e Patient Information 2 e Batch Information 3 Version 1 1 89 of 172 OLI UD 0157 062006 HLATools_ VDManual_v1 1_Rev 0 pdf Short Comments 4 Long Comments Figure 8 14 include MRP count 5 NMDP code definitions for MRP alleles 6 e CR count 7 e CR False Positive and False Negative bead numbers 8 e Review History 9 e Hide Show Report Summary when shown report summary includes user supplied report specific com ments Figure 8 15 10 that are entered in the Report Summary field of the Filter panel Figure 8 10 e Reviewer Information names of sample reviewer and approver with timestamps when two reviewer option 1s in use Figure 8 15 11 e Warning Messages Figure 8 15 12 Rxn Pattern bead NIH scores Figure 8 15 13 e PC Beads Graph Reaction Pattern Histogram Figure 8 16 14 with Positive Control Bead reading Figure 8 16 15 e New Page After creates page break each report prints on a new page Figure 8 14 Sample Report Long Comments Computer Assignment DRB1 0103 DRB1 15HS
86. a code point number to each character in every supported script A Unicode Transformation Format UTF is a way to encode these code points HLATools supports the following options to accommodate different language formats in report generation The ANSI encoding type is the default as explained below You can specify a non default encoding type in the LABType Config uration panel Figure A 4 Version 1 1 109 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf e ANSI the default encoding uses the system s current ANSI code page which is determined by the input language specified in the Regional and Language Options control panel A full listing of code pages can be found at http msdn microsoft com library default asp url library en us intl unicode_81rn asp You can verify which code page your computer is using by selecting Start gt Programs gt One Lambda gt Maintenance gt Connection Diagnostics and finding the value assigned to the CodeSet variable under the Operating System Information section e ASCII encodes Unicode characters as single 7 bit ASCII 7 bit code representing 128 characters and control codes This encoding only supports character values between U 0000 0 and U 007F 127 code page 20127 BIGE encodes Unicode characters in big endian byte order code page 1201 e UTF 16 encodes Unicode characters in little endian byte order code page 1200 e UTF 8 encodes all Unicode characters using 8 bit enc
87. abTypeProduct abTypeProduct 7 R5502Q81_005_0a LabTypeProduct M R55020B1_006_0 LabtypeProduct C Documents and Settings workshop2 1LAMBDAD Refresh Update NMDP gt View This option displays a listing of NMDP allele codes and their subtypes taken from the NMDP Research web site After the first update the revision date of the code listing is displayed above the table Before the first update the legend will state Unable to retrieve This table lists NMDP codes and their numeric allelic subtype definitions The LTI table is initially presented using a numerical subtype sort see inset in Figure 7 8 By clicking on the Code column heading you can re sort the table so that the allele codes are displayed alphabetically The NMDP allele codes are curated by the NMDP Research Program The current version may be found at ht1p www nmdpresearch org HLA numeric html Version 1 1 77 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Figure 7 8 Navigating to an NMDP Code in the NMDP Allele Code List NMDP Revision Date Sunday November 27 2005 NMDP Allele Code List SubType 0212 0225 0227 0231 0232N 0239 0253N 0260 ADXN 0213 0226 0231 0232N 0239 0253N 0260 Code ADRY ADXP 19 36 desd Code SubType EN EII e CU CT oe IE cc CS maw oeseaseneso E E GM 01 02 03 04 05 apxw 0244 0253M 0260 CN AVK 01 02 03 04 05 06 ADX 0248 0249 0253N 0260 om i ADH 01 02 03 0
88. ample s row 1 2 3 10 11 File header panel the same information seen when importing the batch using the Lambda Explorer User identifier computer s network domain and name of currently logged on user Filter options to filter the entire batch according to allele occurrences in various ethnic populations If a sample contains an allele that is not present in the selected population allele assignments are not made for the sample Of the Japanese demographic filters Rank A filter is the most exclusionary and Rank ABC is the most inclusive These population filters can also be applied separately to individual samples in the Sample Assignment Subview right hand panel in Figure 5 8 Batch comments annotations and comments that pertain to the entire batch Unlock All makes all samples in the batch available for editing and changing allele assignments Auto Accept locks all samples in the batch globally This avoids the need to Accept the assignments for each sample individually Locking unlocking individual samples is done in the Sample Assignments Subview right hand panel in Figure 5 8 You can force LTI to reanalyze the entire batch by clicking Auto Accept then Unlock All Note however that this will undo any acceptances that you have entered for individual samples Print outputs a minimal summary report consisting of Sample ID Patient ID well location allele assignments the first line of sample
89. ank 3 Allele Subtypes display of all the alleles which define the code in the Allele column In the case of an unambig uous allele this will simply be the allele itself 4 Search text boxes used to cluster allele reaction patterns separately for Allelel and Allele2 See Sorting by Allele Pattern Matches p 63 and Reaction Grid How is the sorted p 126 5 lt gt swaps the entries in the two search text boxes 6 Go redoes the clustering after an edit has been made to the entries in the search text boxes 7 Bead IDs you can restore the original bead order by clicking in this cell Figure 5 21 Restoring Bead Sort Order 8 Allele lists all the allele specificities in the product alleles with identical reaction patterns are represented by NMDP or local ambiguity codes the code definitions are shown in the right most column of the grid 15 Double clicking on any allele moves it above the Rxn row its final position is determined by its similarity to the Version 1 1 59 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf 10 11 12 13 14 15 Rxn according to the criteria discussed in the FAQ topic Reaction Grid How is the sorted p 126 double clicking on the Allele heading restores the original order with the sample reaction pattern Rxn at the top Rxn sample reaction pattern i e all the observed positive reactions in the sample clicking in the left most cell
90. arded B 44AXJR B 57CG Allele 1 Allele 2 Allele 1 Allele 2 Allele 1 Allele 2 B 4403 B 5703 B 4403 B 5703 B 4403 B 5703 B 4403 B 5703 B 4403 B 5707 B 4403 B 5707 B 4426 B 5703 B 4426 B 5703 B 4426 B 4426 B 5703 B 4426 B 5707 B 4426 B 5707 B 5707 B 4430 B 5707 B 4430 B 5707 B 4430 B 4435 B 5703 B 4435 B 5703 B 4435 B 4435 B 5703 B 4435 B 5707 B 4435 B 5707 B 4436 B 5703 B 4436 B 5703 B 4436 B 4436 B 5703 B 4436 B 5707 B 4436 B 5707 B 4437 B 5707 B 4437 B 5707 B 4437 B 4438 B 5703 B 4438 B 5703 B 4438 B 4438 B 5703 B 4438 B 5707 B 4438 B 5707 B 4439 B 5703 B 4439 B 5703 B 4439 B 4439 B 5703 B 4439 B 5707 B 4439 B 5707 As a final step LTI assigns NMDP codes to all alleles that share the same reaction pattern If the assignment is unambiguous i e the candidate alleles contain no false reactions and are of the same type the alleles in each table are again represented by a more inclusive NMDP code and this assignment is displayed in the column header If a single code assignment cannot be made because the alleles are of different types or they contain a false reaction the typing is considered ambiguous and is denoted by an orange background in the Type Subtype subview Version 1 1 125 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Figure B 10 Type Subtype Tables B S7CG Allele E Filter mo ror B 4437 B 4438 B 4439 sy 35850 adAxqnsfad 4 II It PaYNeEW
91. atically rescales the Y axis to accommodate the range of values in the batch In Figure 5 5 the range of the Y axis has been scaled down to about 2000 2 Control Probe Symbols the display differs according to the product being used in the analysis Class I products employ two positive control probes one for exon 2 and another for exon 3 Class II products employ a single posi tive control probe for exon 2 Green circles represent Exon 3 positive control values These are used only in Class I products Blue squares represent Exon 2 positive control values These are used in both Class I and Class II products See the Glossary for a brief discussion of Class I and Class II MHC proteins The minimum Positive Control Value can be set globally in the HLATools Product Information table Figure 7 3 The default minimum value is 1000 3 Tool tip mousing over a symbol displays a tooltip containing the name value and sample ID of the positive control bead s on the Positive Control Values chart Figure 5 5 Control Values Positive Control Values Positive Control Values 1800 00 1500 00 o e a o 1200 00 No T Minimum Value es o PosCtrl1 600 00 4 Sample B6 1197 00 300 00 a 0 00 T T I i N e9 LO En m 22 co co co co co co co co 2 v 2 v v D v v pat a a a a a a a E E E 0 Q 0 0 0 0 mo hu w w un w wo w wo ow 44 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1
92. aunch the HLATools Configuration Suite and select Products gt View to access the HLATools Product Information table The newly updated product will appear in the bottom row For details on this table see HLATools Product Information p 73 76 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Important Note When you Save a file please note the extension associated with the saved file The internet browser security settings on some computers may suppress the LabTypeProduct file extension when you download the file although the extension does appear when you navigate to the file on the One Lambda LabTypelnteractive webpage Figure 7 5 When these security settings are in effect the LabTypeProduct file extension is replaced with a generic doc extension with the result that your operating system cannot associate the file with the updating utility and it will not be able to load the file information into the LTI database Save the file to a location on your local computer and add the file extension to the filename When you select the Product gt Update option and browse to the folder containing the new LABType file s the updating utility will be able to recognize the files The utility will update your LTT database with the files you select by checking the checkbox to the left of the filename Figure 7 7 Figure 7 7 RSSO Files with LabTypeProduct Extension 8 HLaToo s Product Update x TF R5502Q081_002_R4 L
93. ched reaction pair 161 MRP detection flowchart 119 MSDE Microsoft SQL Server Desktop Engine 13 20 114 116 HLATools User Manual Index 169 of 172 HLATools_IVDManual_v1 1_Rev 0 pdf MSDE version and product level 115 N N80 K80 C locus allotypes 161 Named Pipes 116 136 Negative control 161 Networking protocol versions 115 NIH Score 157 162 NMDP allele codes code definition 158 examples 162 listing 6 75 revision dates 75 scheduling automatic update 128 Normalization formula 162 Null alleles 163 P Patient Explorer purpose of 67 Patient IDs 67 trivial 31 PBS Phosphate Buffered Saline 163 PCR SSP PCR Sequence Specific Primer 163 pNPP 165 Positive controls 164 Class I and Class II 47 156 Positive reaction defined 164 Probe ID 73 Product Lot deleting from Product Information table 71 retired 73 R Reaction Pattern Histogram in analysis report 98 printing 41 viewing entire width 109 Reanalysis of entire batch 35 Recognition Site 164 Report formats A4 page size 91 template files 91 US Standard page size 91 Reports customizing header information 80 customizing patient report 82 overriding default paper size 90 print and export functions 92 printing and exporting 92 report filters 93 Resolution Limitations data sheet 73 S Sample IDs handling duplicate 31 trivial IDs not matched to patient 31 Scripts all cmd 136 Security mode
94. directly to the Luminex 100 PC s Output folder from within the Lambda Explorer Figure 4 1 and Figure 4 2 4 154 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Glossary This glossary contains terms that may be helpful for beginning users of HLATools LABType Interactive A general glossary of terms in HLA immunology and related fields compiled by One Lambda can be found at http www onelambda com resources glossary asp Glossary Entries A 60 A289 Ratio A measure of the purity of DNA in a sample can be obtained by comparing the ratio of spectrophotometric absorbance of the sample at 260 nm to that of 280 nm Nucleic acid samples have a higher absorbance at 260 nm than at 280 nm while protein samples have a lower absorbance at 260 nm than at 280 nm It has been empirically determined that a pure nucleic acid sample has an A gt 2gy A gt gg ratio of 2 0 while a pure protein sample has a ratio of 0 57 Using this ratio as a guide one can estimate the relative concentration of nucleic acid to protein is in a sample The One Lambda Micro SSP protocol calls for a sample Ay60 A280 ratio of between 1 65 and 1 80 which corresponds to a DNA percentage between 65 to 80 ACD Acid citrate dextrose a blood anticoagulant used in preparing samples for PCR Allele Name In 2002 the WHO Nomenclature Committee for Factors of the HLA System decided that Extensions in cases where the total number of allelic subtypes
95. e from a different server than the one on which you are creating the new LTI database replace local with the IP address of the target server or the server name Again consult your database administrator to determine if you should use the default server instance i e either nothing or a named instance 2 In line 3 you can use the existing LTI database name or change it to any name you wish such as MY_LTI_DB 3 SET TABLE MY_LTI_DB 3 Lines 4 and 5 set the login protocol e The E switch invokes Windows authentication If you want to use Windows authentication leave lines 4 and 5 unchanged e Ifyou want to use SQL authentication comment out line 4 and un comment line 5 4 REM SET LOGIN E 5 SET LOGIN Usa Psecret In this case the user name would be sa and the password secret You can change both of these values as you wish For example for the username password combination myname MyWord lines 4 and 5 would be 4 REM SET LOGIN E 5 SET LOGIN Umyname PMyWord Usernames and passwords follow the usual conventions with the username being case insensitive and the password being case sensitive 4 Inline 8 replace the hard coded value LTI with the environment variable STABLES Version 1 1 137 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf 8 osql S SSERVERS SLOGINS q EXIT DROP DATABASE STABLES 5 Comment out line 10 10 REM Del ProgramFiles Microsoft SQL Server
96. e the functions of the Admin Diagnostics Tool and the Quick Connect Tool are now accessed by the Configuration Suite User Management and Connections utilities respectively Version 1 1 111 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Update Database Why can t I run Users without administrative privileges will discover that the Update and Clean buttons are greyed out in the HLA Tools Configuration Suite gt Database gt Local Create panel This is because ordinary users do not have Write access to the folder where the LTT database resides If you encounter this problem ask an Administrator to log on and run these functions for you LTI database does not exist Why do get a message that This message may be generated if you as an ordinary user have attempted to run Update Database You are able to start the process that stops the SQL server service and detaches the existing database However you cannot complete the Update because you lack Write access to the location where the database resides When an Administrator then attempts to run Update Database the application cannot find the database because you have detached it This problem can be solved by an Administrator using the HLATools Configuration Suite Connections tool to reattach the database Select Start gt Programs gt One Lambda gt HLA Tools Configuration Suite gt Connections Select a Connection from the Connections list this wil
97. e 9 2 e Batch ID e Sample ID e Patient ID e Allele Version 1 1 93 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf e Catalog Lot Press Report The report is automatically displayed in the Reports view If the report is For print use the printer icon in the tool bar many print reports can also be exported e For export only use the envelope icon to the right of the printer icon to access a standard Save File dialog Figure 9 1 Export formats include txt pdf xls and doc file formats Figure 9 1 Report Print and Export Functions Patient Reports 1B Patient Summary Report Filtered by Batch The report contains Allele assignments for all samples associated with each patient in the selected batch allele assignments for samples not associated with a patient are also included NMDP codes are expanded to four digits plus N or L suffix Analysis date for each sample Corrected accepted allele assignments are printed in a separate column 2A Patient Report A combined sample analysis report of all samples associated with the specified patient Multiple samples in a single batch or samples from different batches may be associated with a single patient The report includes Full patient data Batch product lot information and locus type Sample draw test and analysis dates Batch comments Corrected accepted typing if any Reaction assignment NIH 8 1 single row format Computer assi
98. e IV gt B 4801 Filter Previous Accept Next Sample ID Sample 5 Date 4 7 2005 B 44025 Ayoypads sxy 358501 adAyqnsjed y Sajellw It Payne Assignment Figure 5 12 shows two type subtype listings 1 Unfiltered shows all the allele assignments for the sample The allele assignments or in the case of multiple sub types the NMDP or local codes are displayed in the column headings 2 Filtered applying a demographic filter to the sample may eliminate numerous subtypes The effect of applying the European demographic filter to the original non filtered subtype listing is shown in red 3 B 44ANAW filtering the alleles results in a smaller number of subtypes in Allele 1 and consequently a different NMDP code 4 DNA in each case the allele assignments are displayed in the DNA assignment fields 54 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Closest Reactions Subview If a Matched Reaction Pair MRP can be obtained by reversing the reaction state of a single bead positive to negative or vice versa that reaction pattern is known as a Close Reaction All Close Reactions for a sample are listed in the Closest Reactions Subview Figure 5 13 The Closest Reactions subview shows what would happen if the cutoff values for a bead were changed without actually doing so The two False Reactions that appear in the Matched Reactions subview in Figure 5 8 each involve onl
99. e LTI data base In this case we will use a wildcard to find all the patients Figure 8 5 whose Patient IDs start with the numerals 243613 and later select several for inclusion in our report After entering the Patient ID search string we press the Search button at the bottom of the panel Figure 8 5 Patient Search Options Search Search Sections Paper Size Logo Inbox O Archive Batch ID x Patient ID 243613 y Sample ID 6 Patient s Inclu PatientID Y Gender Y Race Y Catalog ID Allele NMDP Specificity 2436135 2436136 2436137 2436138 2436139 Reports Filters AX A Search sa 4 The search results are displayed in the Filters panel see side tab in Figure 8 5 You can include one or more Patients in a report by checking the Include boxes on the left A number of the most important patient attributes such as Gender Race etc can be displayed in the Filters panel This table is fully sortable You can sort on any column by clicking on the column heading to the left of the filter cone icon and you can perform a subsort by shift clicking on a second column 5 The Patients selected in the Filters table automatically appear in the Sample s to Report On panel immediately below The default sorting order for this table is by well position although you can sort the samples by the contents of any column in the table In Figure 8 6 the two samples have a low bead count so
100. e Reactions 49 Assignments View w MRPs 45 Attach Detach Database Tool 130 131 Backup Database 23 Batch Results Table 36 Bead Analysis View 39 Bead information where to find 72 Character Encoding Type specifying 150 Class I and Class II MHC Molecules 157 Clean DataBase Log Message 17 Close reaction example 158 Close reactions sorting options 106 Closest Reactions Tables 53 Comments Editor utility 108 Connections Tool 142 Control Values Minimum Bead Count 43 Control Values Positive Control Values 42 Custom Reports Advanced Search Filters 85 Column Search Filters 84 Patient Reports search options 82 Patient Search Options 83 Samples to Report On 84 Customer Information Dialog 15 Database authorized users 141 Database settings 143 Database settings creating 143 Export Type specifying 150 Flowchart Updating HLATools 21 HLATools data reference files 70 Installshield Modify Message 14 LABType Configuration Panel 109 LABType Product Information 71 Lambda Explorer Adding a Folder 28 File Tree 28 Local server reactivating connection to 145 Log on type specifying 150 LTI Batch Sample File Tree 34 Luminex Files folder specifying 150 Microsoft NET Framework Installation 14 MRP tabulation 123 MRPs from Golden alleles 121 NMDP code definitions where to find 76 Patient Information Subview 55 Patient Typings Table 68 Probe recognition sit
101. e between tabbed windows e Select any tab in a set of tabbed windows to establish the focus on that set of tabs Thus to navigate in the batch windows select either the Batch Results Bead Analysis or Control Values tab e Select ctrl tab to navigate to the next tab in series e Select shift ctrl tab to navigate to the previous tab in series e Press enter to access the window Accessing LTI Functions via Hot Keys After you launch LTI press the alt key to enable the hot keys The hot key characters for the function will be under lined on the appropriate control You can execute the function by using the alt key combination Changing Date Formats in HLATools The date format is determined by Windows system settings To change the date format in Windows XP e Select Start gt Settings gt Control Panel gt Regional and Language Options In the Regional Options tabbed window select the desired region from the pull down list HLATools will use the date format displayed in the Short date text field To change the date format in Windows 2000 e Select Start gt Settings gt Control Panel gt Regional Options e In the Date tabbed window select the desired date format from the Short date format pull down list Editing the Assignment Subview Comments List The Assignment Subview contains a pull down list which contains frequently used stock phrases that you can add to sample specific comments Any user defined comment
102. e different positive controls Class II products are assays for only a single exon If a positive control is low a warning appears in the Reaction Pattern Histogram for the sample Figure 5 8 A bead reaction that has been changed due to a user modification in the cutoff value is also designated in the raw data table by a light purple background color Figure 5 28 Figure 5 28 Low Readings in Raw Data Table jajaa ad a em pasos o a E 32 8 Posie Cornet 100 1007 140 1 83 pre gt e es In Figure 5 28 the reading for bead 30 is near positive right at the cutoff value of 10 and the value for one of the positive controls is below the default 1000 minimum 66 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Figure 5 29 Raw Data Table Double Column Sort i a Recognition Site seit Cutoff Count NC PC 60 G 65 23 292 q B 570101 e 0520 1232 OTT BF FS a E IE A Ara a E 04220 ea erro Caer armar eiee a 3 2 ap 1 Rxn the table in Figure 5 29 was first sorted by reaction status with negative reactions 1 sorted before positive reactions 8 2 Raw a secondary sort was then made for ascending raw bead readings 3 Bead clicking on the Bead heading restores the table to its original sort order Version 1 1 67 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf This page intentionally left blank Chapters Patient Explorer ee Patient Explorer
103. e selected i e GER_MSDE2000a exe for German or JPN_MSDE2000a exe for Japanese 5 Double click on the file and select a directory to extract the installation files into 6 Select Start gt Programs gt Accessories gt Command Prompt Using the command prompt navigate to the extrac tion directory To install to a default instance enter setup SAPWD AStrongPassword TARGETDIR C MyInstanceFolder To install to a named instance enter setup SAPWD AStrongPassword INSTANCENAME ONELAMBDA TARGETDIR C MyInstanceFolder Note that the directory specified for the TARGETDIR property must always include a trailing back slash How can import LABType Classic analyses into HLATools The LABType Database Upgrade utility allows you to import analysis results from a LABType FoxPro database into an SQL database When you migrate a LABType FoxPro database the installation program checks whether the required Microsoft Net and MSDE software components are present on your system and will install them if necessary In addition the installation program will create an SQL database if none is present These actions are executed automati cally and do not require user intervention To migrate your data from a LABType FoxPro database into an SQL database 1 Locate the Database Upgrade folder on your LABType distribution CD and launch the setup exe inside 2 Follow the InstallShield Wizard instructions accepting the usage agreement ter
104. ears in the Connection Name field Figure C 7 The More button accesses a list of other accessible servers in the pull down list If this list does not populate check that the Named Pipes and TCP IP communications protocols have been enabled For details see Peer to Peer Installa tion p 135 11 Accept the default options for Windows NT Integrated Security 2 and LTI for database selection in 3 Select the desired server from the pull down list Figure C 8 or enter its name directly into the field The name must observe normal pathname naming conventions and not contain control characters or whitespace In this example we will select the server swordfish where we just added the remote user Quimby to the authorized users list Version 1 1 143 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure C 8 Available Servers Connection Name Homer 1 Select or enter a server name swordfish salmon shark skipjack starfish E 12 Press Set and then Test Connection to confirm that the connection has been established 13 Now that the connection Homer to the LTI database on swordfish has been established we must activate the con nection before launching HLATools After selecting Homer from the Connections list choose the Make Active option from the right click menu This activates the Homer connection as shown in Figure C 9 Figure C 9 Activating a Server Connection la File Analysis
105. el gt Add or Remove Pro grams from the Windows desktop or use the removal tool provided with the HLATools Install program Figure 2 5 Install the new version of HLATools as described above in Software Installation p 15 At this point you have to decide whether to update the existing database create a completely new database by run ning Clean Database or maintain the current database unchanged The Readme txt file on the distribution CD contains instructions on which path to follow A Run Update Database when modifications have been made to the database or when the product nomenclature has been updated This preserves the results of your previous analyses e Select Start gt Programs gt One Lambda gt HLATools Configuration Suite gt Clean Update then select Update e The Update utility will take several minutes to run Its progress is displayed in a log message window B Run Clean Database when you want to create a completely new database structure See the Clean Database Glossary topic for details Version 1 1 OLI UD 0157 062006 25 of 172 HLATools_IVDManual_v1 1_Rev 0 pdf Log On In the standard installation of HLATools on a standalone workstation any user can access HLATools without further login Further levels of security can be implemented to control access to the applications The implementation of restricted access is discussed in HLATools Security Modes p 149 Appendix C 26 of 172 6 6 06
106. er s local LTI database the target user must add the remote user to the list of authorized users on the target computer This can be done remotely as well as will be discussed in Editing the Authorized Users List from a Remote Server p 146 In setting up peer to peer access it is necessary to know the network identities full computer names of the computers You can find this by selecting My Computer gt Properties gt Network Identification from the Windows 2000 desktop or My Computer gt Properties gt System Properties gt Computer Name from the Windows XP desktop In this example we establish connections between two servers named swordfish and skipjack 1 On the target computer swordfish launch the User Management tool by selecting Start gt Programs gt One Lambda gt HLATools Configuration Suite gt User Management 2 Select List Users to refresh the list of users who currently have access to the swordfish LTI database Figure C 2 138 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure C 2 User Management Tool HLA Tools Interactive Configuration Suite File Analysis gt swordfish ONELAMBDA Comments II sa Configuration BUILTIN Administrators coomo asst yaari Database swordfish Homer Attach Detach Backup Restore Clean Update Merge Diagnostics User Management List Users Add SQL User Add Windows User Group Delete User 3 Two methods of a
107. erence values from One Lambda product data sheets and then suggests likely batch product lot matches as shown in Figure 4 4 If you don t know the lot information select the first lot in the list If the lot information is displayed in the header panel select that lot If the match is exact Lambda Explorer will proceed to load the Luminex file into the database However if there is a mismatch you may encounter a message like this Figure 4 5 Sample Product Mismatch upon Import Import using RSSO1B Lot 002 rey 0 gt i xi Product does not match The sample you are importing has extra probes not needed by this product Do you still want to import using this Product eT A similar message appears if the samples in the batch have fewer bead types than the product Important Note Please note that neither a too many probes nor too few probes message necessarily means that the analysis cannot continue Laboratories may customize their samples so that they do not exactly match a One Lambda product template or a systematic laboratory error may have the same effect 32 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf After you OK the import or bypass the mismatch message LTI executes five steps when importing a batch file into the HLATools database These are typically Trimmed Mean Count Normalizing Analyzing and Matching Patients Trimmed Mean indicates that LTI is using as raw data t
108. erologically this is indicated by the symbol B or C Print prints the OLI product name batch name and sample specific information contained in Matched All Alle les Type Subtype and Closest Reactions subviews The Reaction Pattern Histogram is printed on a separate page Restore removes any user edits or assignments for the current sample from the database Locked prevents further edits or assignment changes from being made to the sample When this checkbox is grayed out the sample is not locked To lock the sample press Accept which enables the Locked checkbox and checks it A locked state for the sample is also indicated by the entries in the DNA allele assignment fields being displayed in bold type Locked samples are checked in the Batch Results View table Figure 5 3 48 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 11 Filter filters the current sample locally using one of the Japanese or European filters discussed above in the Batch Results View Allele assignments made in filtered samples are indicated by red type in the All Alleles and Type Subtypes subviews 12 Comments a short listing of frequently used user supplied sample comments a comment selected from the list is appended to the sample comments contained in the field below This entries in the list can be edited using the Com ments Editor utility see Editing the Assignment Subview Comments List p 107 13 Comments Tex
109. erted here for reference only 1 echo off 2 SET SERVER local OneLambda 3 SET TABLE LTI 4 SET LOGIN E 5 REM SET LOGIN Usa Psecret 6 REM SET SERVER 192 168 1 200 OneLambda 7 net start mssqlS onelambda 8 osql S SERVER S LOGIN q EXIT DROP DATABASE LTI 9 net stop mssqlS onelambda 10 Del SProgramFiles Microsoft SQL Server MSSQLSONELAMBDA Data LTI 11 net start mssqlS onelambda 12 osql S SSERVER LOGIN q EXIT CREATE DATABASE LTI 1 Inline 2 local OneLambda defines the server name and server instance Note that a server instance does not need to be defined in which case the database will be created in the default server instance Alternatively you can specify an existing instance e Ifyou intend to run all cmd locally i e from the same server in which you are creating the new LTI database leave local unchanged The database administrator may want the LTT database to reside on the default server instance of the server to reduce resource usage The default may either be unnamed i e null or named In this case delete OneLambda and replace it with the default either nothing or with the name of the default server instance Otherwise the script will create a separate OneLambda server instance e Ifyou intend to run all cmd remotely i
110. es 165 Product Information Controls 70 Product Update downloading LABType product information 74 missing file extension 75 Product Lot Selection 30 Program Maintenance 18 Raw Data double column sort 65 low bead count 64 low readings 64 sort by minimum bead count 64 168 of 172 HLATools User Manual Index 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf sort by recognition site 63 Reaction Grid Golden allele 121 bidirectional sort by allele pattern 62 clustering positive reactions 60 default view 120 False Negative 59 False Positive 59 restoring allele sort order 61 restoring bead sort order 60 sort fields 61 sorting by bead usage 59 Rearranging table columns 106 Report header customizing company icon 81 laboratory information 81 Report Header Laboratory Information 80 Report Templates folder specifying 150 Runtime Error Message 19 Sample Reaction Pattern Grid 56 Sample Product Mismatch 30 Selecting a File for Import 29 Serological equivalents where to find 77 Server activating connection to 144 Server listing available s 144 Server listing local users 146 Server Quick Connection to remote 147 Server connection activating 144 Setting Database Name in ALL bat 131 Setup Options Custom 19 Simple Network Installation 139 Specificity Subview 54 SQL authentication 148 SQL Server and Database Status 22 SQL Server Netwo
111. exceeds 99 a second number series would be used to the extend the first one This nomenclature has been now been imple mented for several alleles the extension of the B 15 type or family is B 95 and the extension of the A 02 type is A 92 These nomenclature extensions are observed by LTI See http www ebi ac uk imgt hla nomen_changes html Amino Acid Codes The following Amino Acid Codes are recognized by the International Union of Biochem istry IUB and the International Union of Physical and Applied Chemistry IUPAC Version 1 1 HLATools User Manual 155 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Table Gl 1 IUB IUPAC Amino Acid Codes Code Three Letter Abbreviation Description A Ala Alanine B Asp Asn Aspartic Acid Asparagine Cc Cys Cysteine D Asp Aspartic Acid E Glu Glutamic Acid F Phe Phenylalanine G Gly Glycine H His Histidine lle Isoleucine K Lys Lysine L Leu Leucine M Met Methionine N Asn Asparagine P Pro Proline Q Gin Glutamine R Arg Arginine S Ser Serine T Thr Threonine V Val Valine Ww Typ Tryptophan X XXX Unknown Y Tyr Tyrosine Z Glu Gin Glutamic Acid Glutamine K TERM Termination Stop Codon ASO Allele specific oligonucleotide typing also called sequence specific oligonucleotide SSO typing Class I MHC Class I MHC molecules usually display endogenous proteins or peptides derived fr
112. ff 10 2436181 2436162 2436199 2436149 2436219 2436223 2436141 2436136 2436217 2436212 2436186 2436161 2436178 2436158 2436198 2436202 2436135 2436140 2436112 13 I False Positive Ml False Negative Jump directly to a bead by E double clicking on the Bead ID Print Pie ON LOMA Or OSLO Pa Or 20 000 CLO Pe 9 OSLO Pe 00 09 LOOP 00 09 LO 0 OSLO 00 DON LO Mr OSLO Vv rer A OS OSOS OSOS 09009 09 09st e a LO LO LO LO LO LOLO LOLO Cor Fr 09009 0900 0005 00 v Locked Cer LO 000 Oe 1000 DON LO MaN 09 1000 ODO LO 00 Ory LO OOO Ce 1000 O Ce M 1000 O Ot AA A OS OS OS 09009 09009 eet eet LOLO LO LO LOCO LOCO LOLO Pa PP Pa 0009 0900 09 030 Batch Results Bead Analysis Control Values Version 1 1 41 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Bead Analysis View The QC Data and Sample Data Bead charts allow you to observe on a bead by bead basis how the batch samples compare to a representative cross section of the general population In both charts the bead readings for all samples are displayed with values increasing from left to right Clicking on the thumbnails 4 5 displays full sized versions of the charts in the main panel The False Reaction Indicator 13 in the bottom panel does not change when you toggle between the QC Data 4 and Sample Data Bead 5 charts Table 5 1 Bead Analysis Color Codes Reading Color Description More than 110 of cutoff assuming default 10 c
113. fication number 2 Specificities complete currently known specificities for the bead 3 Mouse over specificities appear when mousing over the bead in the Reaction Pattern Histogram at the bottom of the Assignments view Figure 5 8 4 Right click menu text field contents can be copied to the clipboard This is a non editable field Sample Assignment Subview This subview and its features were discussed above together with the Assignments View and the Matched Reaction Pairs subview Please see notes to Assignments View with Matched Reaction Pairs and Close Reactions p 47 56 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Patient Information Subview This subview is where the reviewer can enter patient information Note that multiple samples may be associated with one Patient Information entered in the Patient Information Subview applies to all samples for a patient Sample information pertains only to the sample All entry fields are optional and most require no explanation The entry fields are not locked until either the Match or Save operations have been carried out Figure 5 15 Patient Information Subview Sample Sample 6 m Patient Information 4 ID Hamilton 2 Last Hamiton Fist Bob sts Midde 3 DOB 01 01 1935 Race Eng Gender M ABO RH Run with Euro filted 5 Match Save 6 Locked Mach seve 5 Relationship Matemal half brother Family 8
114. formation and locus type e Test date for each sample e csv filepath e Batch comments are omitted e Corrected accepted typing if any for each sample e Reaction assignment NIH 8 1 format e Computer assignment combining all MRP designations into a single NMDP code pair if applicable e Generic allele descriptions with complete listing of types and subtypes for each MRP e Listing of close reactions Version 1 1 97 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf e Expansion of NMDP codes for computer allele assignments 3G Combined Batch Report Sorted by Patient This report contains data for each patient assembled from each batch in which an associated patient sample appears Draw dates for the samples are not included The patient information for each patient includes Patient ID name gender birthdate race and blood typings e Batch product lot information and locus type e Test date for each sample e csv filepath e Batch comments are omitted e Corrected accepted typing if any for each sample e Reaction assignment NIH 8 1 format e Computer assignment combining all MRP designations into a single NMDP code pair if applicable e Generic allele descriptions with complete listing of types and subtypes for each MRP e Listing of close reactions e Expansion of NMDP codes for computer allele assignments Sample Reports 3C Sample Report Filtered by Batch This brief report contains data for each sa
115. ften corresponds to an Allele1 or Allele2 match while one with a high density in the remainder of the pattern corresponds to an Allele2 or Allele1 match Figure 5 24 shows a small section of the reaction grid for a B locus sample Due to space constraints many rows above and below those shown have been omitted Similarly a large number of columns have been omitted Many of the omitted columns contained varying numbers of false negative reactions Note how this bidirectional sorting forces the alleles with high proportions of positives at the beginning of the sample reaction pattern to cluster above the Rxn row and those with positives in the remainder to cluster right below it This clustering makes it easier to visualize how various combinations of alleles can yield Matched Reaction Pairs MRPs and Close Reactions In this particular sample the junction of B 1502 Allele2 and B 0723 Allele1 yields one of several MRPs Raw Data Table The raw data table contains bead specific raw and normalized data All columns in the table can be sorted alphanumer ically by clicking on a column header A secondary sort can be made by shift clicking on a second column header Clicking on the Bead column heading restores the original sort order For an example of a secondary sort see Figure 5 29 The following notes refer to Figures 5 25 through 5 29 64 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf e Bead bead ID numbe
116. gnment combining all MRP designations into a single NMDP code pair if applicable Generic allele descriptions with complete listing of types and subtypes for each MRP Listing of close reactions Expansion of NMDP codes for computer allele assignments 94 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Figure 9 2 Report Filter Interface m Report Filter Batch ID 4ug04WorkshopDRB1LocusTest SampleID Sample5 o Patient ID SSEEEE y Allele Catalog Lot Lot Rev Report Reports Summary Information 44 Sample Raw Data Abbreviated Specificity Report Filtered by Batch One Lambda Inc Generate the report by selecting a batch and then selecting a sample from withing the batch The report will include only sample and the corresponding analysis from the batch selected Filters Designer Batch Reports 3A Batch Report This report format includes as many samples as possible on each page No patient data Batch product lot information and locus type Test and analysis dates for each sample csv filepath Batch comments Corrected accepted typing if any for each sample Reaction assignment NIH 8 1 single row format Computer assignment combining all MRP designations into a single NMDP code pair if applicable Generic allele descriptions with complete listing of types and subtypes for each MRP Version 1 1 OLI UD 0157 062006 95 of 172 HLATo
117. gt gt TCP IP lt lt Disable Force protocol encryption Properties Tl Enable WinSock proxy Winsock proxy address Winsock proxy port 3 Select the ONELAMBDA instance of the SQL server installation on the local computer in this case swordfish 4 Make sure that both Named Pipes and TCP IP have been moved to the Enabled protocols field OK the newly enabled protocols Any changes that have been made are thus saved but will not go into effect until the computer has been restarted 5 Repeat this procedure on every computer that will be sharing LTI databases Creating a Database on a Remote Server The all cmd script is located in the C Program Files One Lambda HLATools Clean Database folder This script is launched when you run the Clean Database Utility which creates an LTI database on your local computer see Running the Clean Database Utility p 18 You can use this same script to create an LTI database on a remote computer Of course you must have write access to the remote computer to do this To create a database on a remote computer you will need to edit the server name the server instance name and the database name in the all cmd script These changes involve only the first few lines of the file 136 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf When you open all cmd in a text editor the pertinent lines will appear as follows without the line numbers which are ins
118. gure 5 1 Connections Tool Provides access to multiple LTI databases on networked servers For details see Connecting to a Remote LTI Database p 138 Database Tools Attach Detach makes or breaks connections to LTI databases For details about the need to detach databases see the FAQ on excessive growth in database size Appendix B p 129 Backup Restore displays the current instance of the SQL server and the attached LTI database and backs up an ex isting LTI database or restores a backed up database located on a local or a remote server Clean Update creates a clean or new LTI database or updates an existing one See the Clean Database and Up date Database Glossary topics Merge combines two existing LTI databases See the Merge Database Glossary topic System Diagnostics Displays information about e current user and users authorized to use local computer e local computer OS hardware components and IP address e creation dates and last time of use of program components e system environment variables e currently implemented LTI database connections Version 1 1 9 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf User Management Creates new users authorized to access a local or remote SQL database Adds Windows User and Groups authorized to access databases For details see Figure C 2 User Management Tool Features The HLATools software suite is designed to be used with One Lambda
119. he Assignments view These subviews display information about the sample and also contain tools and fields for analyzing and modifying the sample allele assignments Figures 5 8 and 5 9 display the Matched Reactions subview the Sample Assignment subview the Control Values panel and the Reaction Pattern Histogram Figure 5 8 shows a typical sample with two Matched Reaction Pairs while Figure 5 9 shows a sample with two Close Reactions The following notes apply to both figures Figure 5 8 Assignments View with two Matched Reaction Pairs Serological TA jess o Restore i 0 jE Locked Filter Unfitered y Comments Matched Reaction Pairs 3 DNA B 184ETB B 524D B 180101 B 520101 B 1805 B 1817N 5 A E adfiqns ad Low Bead Count 2 Matched Reaction Pairs B 184ETB amp B 524D gt B 180101 B 1805 B 1817N amp B 520103 B 5204 B 184ET8 amp B 520101 gt B 180101 B 1805 A4 1817N amp R52MM1 14 Previous Accept Next 38 15 Sample ID 24361 Date 6 24 2005 Birth 1 1 1901 16 Race Unk Assignment Patient 17 BM SAETB B 520101 SUK 5950 7 Ayoyisads 22 LULA A UA A ULA UA Weal naa eee TNO tT LOW 0 Or NT LO LOA OM Or N SLO LO 00 MO SOON OOO OS mr prr ee ON Assignments Bead Analysis Asn Grid Raw Data 29 Version 1 1 47 of 172 OLI UD 0157 062006 HLATo
120. he Luminex LABScan lt SuperScript gt 100 csv output files which are the input files for the LTI application Files are displayed as follows Figure 4 1 e Files in bold face type have not yet been imported into the HLATools database e Files in regular type have already been imported Attempting to re import a file already in the database will gener ate a warning message Lambda Explorer parses the files to assure that their formatting is correct Corrupt files are identified with a trash can icon e g 041204BL62 csv in Figure 4 1 Corrupt files cannot be imported in LTI Version 1 1 29 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Figure 4 1 Lambda Explorer File Tree 3 LuminesFiles Eg 041204BL62 csv _ 08 12 04 PP1356 DPB1 150 194 CODED csw 03 03 04 RSSO1BJ_001e Final QC conv CS _ 10 08 04 PP1369B LOT7 FINAL 11P AM CODED csv i BL6LowFirstS ample csv 1 BLOCUS lot6 csv 123_104B_L6_ID1473 csv ligh_DR8_pll_96 csv To import files from a non default folder use the browse function by right clicking anywhere in the file tree panel Figure 4 2 and select Add Folder A conventional Find File dialog box appears Figure 4 2 Lambda Explorer Adding a Folder gt A PP1356 DPB1 150 194 CODE Delete RSSO1BJ_O01e Final OC conv 10 08 04 PP1369B LOT 7 FINAL1IPA BL6LowFirstS ample csv BLOCUS lot6 csv 123_104B_L6_ID1473 csv ligh_DR8_plil_96 cs
121. he Reaction Grid header panel contains two text entry fields that are used to enter allele designations LTI then sorts all the alleles in the reaction grid so that you can inspect the extent to which the specified alleles match the sample reaction pattern In the following discussion the alleles specified in the fields will be referred to as First Allele and Second Allele Figure 5 23 Reaction Grid Sort Fields Print B O7Xx1 7 B 1 5X2 Pin Entering a First Allele in the left hand sort field sorts and clusters the alleles according to the degree that their patterns match that of the First Allele Figure 5 24 In the first analysis the criterion for matching the First Allele is that the alleles share at least 75 of the positive reactions of the First Allele The order in which the alleles are sorted is deter mined by this criterion and four other scores for each allele The scoring and sorting protocol is discussed at length in the FAQ topic Reaction Grid How is the sorted p 126 The alleles considered similar are sorted upwards from the specified allele in terms of decreasing similarity with the most similar allele being placed directly above the First Allele and the allele with only 75 similarity ending up at the top of the grid The sample reaction pattern appears directly below the First Allele At this point the remaining alleles in the grid are ordered in terms of their similarity to the allele directly below the
122. he time of the Luminex run Aug04Workshop This ID is the only user editable field in this panel and allows the user to modify the Session ID at the time of file import Double clicking on the ID enables an edit field Edits to the Session ID affect only the version of the file imported into the database and not the original input file thus preserving the integrity of the original data 5 Product designation One Lambda product designation RSSO1A_005 assigned at the time of the Luminex run Note the product designation as it will help you select the proper product lot during the next step of the import pro cess 6 Lot or template name One Lambda lot name assigned at the time of the Luminex run LabType A Locus Lot 005 Version 1 1 31 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf 7 Import saves any edits to the Session ID and brings up the Product Select dialog box Figure 4 4 Figure 4 4 Product Lot Selection Product Select Select matching One LambdaAssay used with this Batch 1O x Augl4 Workshop4LocusT est 7 29 2004 12 09 28 PM ASSO1A4 Lot 005 rev Da Product RSSO1A Lot 004 rev Od ASS5O14 Lot 005 rev Da RSSO1A Lot 006 rev 0 ASS02041 Lot 001 rev O AS50154 Lot 001 rev Oc RASSO1AJ Lot 002 rey O RSSO2Q61 Lot 003 rev Od RSSO2QB1 Lot 002 rev R2 When importing Luminex output files Lambda Explorer compares bead information in the samples with ref
123. he values in the Luminex data file in which the upper and lower 5 of the readings for each bead have been discarded before the mean is calculated The use of Trimmed Mean data is the default setting for LTI Other data types such as simple Mean or Median or Trimmed Median can be specified by the user in the HLATools Product Information Table which can be accessed by using the HLATools Maintenance Tool Figure 7 3 This datatype specification is made on a product lot basis Count imports the bead count for each bead type at each sample well A Trimmed Count option is also available in which the count for each bead is reduced by a specified percentage The default value for this reduction is 10 The count type can be specified in the HLATools Product Information Table Normalizing normalizes the specified mean or median value for each bead the Normalization formula can be found in the Glossary Analyzing determines which pairs of alleles recognized by the LABType product have combined reaction pat terns consistent with the sample reaction and uses this information to construct the Reaction Pattern Histogram for the sample Matching Patients matches samples to patients in the database If a Patient ID is not associated with a sample the sample ID will serve as a default patient ID Thus if a sample in a newly imported batch has the same sample ID as a sample already in the database and the latter sample is associated with a patient
124. hen allelic subtypes are included in a range only the first four digits in the designation are significant all digits signifying synonymous intron and null or low expression polymorphisms are disre garded Thus B 1542 44 could also be the abbreviation for the series B 1542 B 154201 B 15420101 B 154202 B 1543N and B 1544 Because information is lost by abbreviating the specificities LTI uses the complete specificity listing when matching allele pairs with the observed sample reaction pattern For a discussion of allele designation see Alleles How are they designated in Appendix B e RecognitionSite probe recognition site s these entries describe the site s on the locus that are recognized by the probe on a given bead For details on how to interpret these entries see the Recognition Site glossary topic e ProbeID OLI probe identifier e CutOff the minimum value of the normalized bead reaction that is considered positive editing this value for a particular bead will change the cutoff in all analyses of batches using this product e CompleteSpecificity a complete listing of all allelic subtypes to which the probe sequence on the bead will bind Products gt Update The HLATools installation includes a utility that lets you update the LABType product information used by LTI This information is the same as that contained in the Bead Probe Information and Resolution Limitations data sheets for a given LABType produc
125. hes for allele pairs that are two aways three aways and so forth This search mode can be invoked by changing bead cutoff values globally in the Bead Analysis View or locally in a sample s Reaction Pattern Histogram HLATools User Manual 159 of 172 HLATools_IVDManual_v1 1_Rev 0 pdf Genetic Code Immunomagnetic beads JPN Rank Local Code Log Files ldf The amino acids are encoded by the following nucleotide triplets See Table Gl 1for the full names of the amino acids Table Gl 3 Amino Acid Nucleotide Triplets T Cc A G Phe F Tyr Y Cys C T Ser S Leu L Term Term Trp W His H c Leu L Pro P Arg R Gin Q Asn N Ser S lle 1 A Thr T Met M Lys K Arg R Asp D G Val V Ala A Gly G Glu E O D O AO0 P JO AO PB O AJO PO A Immunomagnetic beads are superparamagnetic particles with monoclonal antibodies coupled to their surface The beads can be collected using a magnetic field When the field is removed the beads do not retain any residual magnetism which is one property of paramagnetism the term superparamagnetism indicates that the beads display paramag netic behavior at a lower temperature than would normally be expected superparamagnetism tends to become more pronounced as the size of an object dimin ishes They can be repeatedly magnetized and
126. hus 1f the cutoff for Bead 17 in Figure 5 13 were changed to force the reaction to be negative the allele assignments would be those shown in the table If a cell in either of the Allele columns displays an NMDP or local code selecting the cell causes its definition to appear in the Listing to the right 1 2 Pull down list definition of the NMDP or local code 4 and low expression alleles Version 1 1 OLI UD 0157 062006 Listing definition of the NMDP or local code including synonymous and intron polymorphisms and non coding 55 of 172 HLATools_IVDManual_v1 1_ Rev 0 pdf 5 Collapse Expand closes opens all false reaction subtables simultaneously Individual subtables can be opened closed by toggling the sign to the left of the subtable heading Figure 5 13 contains eight subtables Specificity Subview The Specificity subview Figure 5 14 displays the complete allele specificities for each bead in the sample Figure 5 14 Specificity Subview DRB1 010101 DRB1 010102 DRB1 010201 DRB1 0104 DRB1 0105 DRB1 0107 DRB1 0108 DRB1 0111 DRB1 0404 DRB1 040501 DRB1 040502 DRB1 040503 DRB1 040504 DRB1 0408 DRB1 0410 DRB1 0419 DRB1 0423 DRB1 0428 DRB1 0429 DR81 0430 DRB 0440 Delete Select All Aypypads SUKY 5950 5 adAjgnsfadA sajajjy v payzqew it UA AAA A ESI Se ae a O 1 Bead ID bead ID number in the batch not the One Lambda bead identi
127. ically ends with the locus that is ana lyzed by the product You can delete a product from the table by clicking immediately to the left of the ProductName to select the entire row and then pressing lt Delete gt This action cannot be undone QABatchID the database batch ID number for the product lot Loci the locus analyzed by the product Lot the product lot number product run Revision revision number of the source sheet from which the product information was taken DataType the entry in this editable field instructs the software which raw data from the Luminex output file to use for normalizing the bead readings Trimmed Mean is the default For details and examples see the Mean and Median Glossary topic CountType the entry in this editable field instructs the software which count data from the output file to use The full count is the default See the Count Type Glossary topic Version 1 1 73 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf MinBeadCount a different minimum value can be set for each product lot For example if a product lot contains a smaller number of one or more beads than normal the default is 100 this value can be adjusted accordingly to prevent the samples from being rejected during analysis The value for MinBeadCount can also be changed global ly using the Tools gt Global Settings menu option MinPCData the minimum Positive Control value The default is 1000
128. ides access to several kinds of information Mousing over the bead ID number displays a tooltip with information about the bead reading Beads 51 and 9 are examples of positive and negative reactions respectively e Clicking on a bead ID number re sorts the Reaction Grid so that all positives for that bead move to the top of the table and also displays the allele specificities for the bead in the header area Figure 5 19 shows the alleles in which bead 51 is known to display a positive reaction Allele Code Definition not shown the allele code definition for an NMDP or local ambiguity code is repeated in the right most column of the grid 60 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 16 Print prints out the reaction grid as currently displayed with definitions for the alleles on the right due to its width the reaction grid should be printed out in landscape orientation preferably on at least 11 x 17 inch paper Figure 5 17 Reaction Grid False Positive Allele DRB1 1105 DRB1 1404 DRB1 1428 DRB1 1431 DRB1 1411 DRB1 0809 DRB11415 DRB1 0821 DRB1 1205 Sorting the Reaction Grid LTT incorporates a number of very useful sorting functions that simplify comparison and inspection of alleles and reaction results in the Reaction Grid When you first access the Reaction Grid the observed sample reaction profile and the Golden alleles appear at the top Figure 5 16
129. iew p 53 and Sample Assignment Subview p 56 Analyze sample reaction patterns in a serogram like grid Reaction Grid p 58 Review raw data in a tabular format Raw Data Table p 64 Manage data from multiple samples associated with a single patient Chapter 6 Patient Explorer Update the data resource files used by LabType Interactive Chapter 7 Reference Files and Their Maintenance Generate a wide variety of sample patient and batch reports Chapter 8 HLATools Lambda Reporter and Chapter 9 HLATools Legacy Reporter 10 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf About this Manual This manual is intended to provide the basic information a user needs to take advantage of the advanced features of LABType Interactive and the other modules in the HLATools software suite and to understand the purpose of each element contained in the module interfaces It is assumed that the reader has read and understood one of One Lambda s LABType SSO Product Inserts and is familiar with the basics of PCR amplification and data collection using the Luminex LABScan lt SuperScript gt 100 Flow Analyzer A number of terms pertinent to LTI and other HLATools modules and to other One Lambda products are contained in the Glossary for easy reference Details about laboratory aspects of HLA typing LabType assays or statistical analysis are beyond the scope of this manual How to Use This Guide This guide is d
130. ility Click Next to proceed 5 The name of the selected application will automatically populate the task name text entry field You can rename the task if you want to 6 Specify a Start Time and Start Date For performance frequency select Weekdays since the codes are updated daily Monday through Friday by NMDP Research Click Next to proceed 7 An authorization dialog appears next Your username or that of the current user if someone other than you is logged on to the computer appears in the first text field Enter and confirm your Windows log on password Click Next to proceed 128 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 8 If the password matches the username Windows will schedule the automatic task as shown in Figure B 11 This listing also lists when the last time the utility was run and when it is next scheduled to run Click Finish to com plete the automatic scheduling You can modify schedule settings by double clicking on the scheduled task in the listing Figure B 11 Schedule Tasks Updating NMDP Codes Automatically alo x File Edit view Favorites Tools Advanced Help ay yy Bact an gt T JO search Folders 3 Es x E Address E scheduled Task xf Go Name A Schedule Next Run Time Last Run Time Status Last Result Creator 2 Add Scheduled Task Pua Update NMDP Data At 1 02PM 1 02 00 PM 10 4 2005 1 02 00 PM OxcOO00 wwwhite 1
131. in the Programs and Services field for tcp and udp Figure B 1 the firewall is blocking access to the remote database 112 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 2 First add a tcp port In the Programs and Services options check File and Printer Sharing then click Add Port 3 Inthe Add a Port dialog select the TCP option and enter the following values Name 1433 tcp Port number 1433 OK the entries The Programs and Services field will now contain a new entry 1433 tcp 4 Next add a udp port Click Add Port a second time In the Add a Port dialog select the UDP option and enter the following values Name 1434 udp Port number 1434 OK the entries The Programs and Services field should now appear similar to Figure B 1 Figure B 1 Modifying Windows XP Firewall Settings Windows Firewall x General Exceptions Advanced Windows Firewall is blocking incoming network connections except for the programs and services selected below Adding exceptions allows some programs to work better but might increase your security risk Programs and Services Name Y 1433 tcp 1434 udp File and Printer Sharing Remote Assistance Remote Desktop UPnP Framework Add Program Add Port Edit Delete V Display a notification when Windows Firewall blocks a program What are the risks of allowing exceptions Version 1 1 113 of 172 OLI UD 015
132. ing any unsaved changes Restore restores the columns and column order to the HLATools default 123_104B_L6 11 2 2004 5 22 22 PM Figure 5 3 Batch Results View Filter Q Sool ma lucy C JPN Rank AB e 36 C JPN Rank A Auto Accept C Europe Catalog ID ASSO1B Lot 006 rev De None 3 Print 1LAMBDA W White 2 BS 9 BatchResultsGrid Sample ID Well Allele 1 ps Allele 2 Comment Patient ID Lek Relations A 2436151 18 B3 2 emma mr g BatchID 4 y CR y MRP v Comment II v NMDP 1 v NMDP 2 v Al 4Digit v A2 4Digit ZI 3 w me w nO 4 Batch Results Bead Analysis Row color codes B 404UC B 5801 Low Bead Count 2436151 F rer ee eae AFJY Low Bead Count 2436166 O 2 v Sample ID cod e Serological K biguity B44 B12 B44 B12 Restore e Serological Abig 2436204 a B aanvu B 57GW Low Bead Count 2436135 dul Half Family Al Az Sero 1 Sero2 Bw al B 40 B 58 a a fas Ae fees pp fewsfewfifi ED B44 857 3l3 B9 657 bs 657 Bw4 Bw4j2 1 EE ABE Ce eee PA Cese ese J o J A Pee S gt A Control Values e Blue White Matched Reaction Pairs MRPs have been assigned i e the combined reaction patterns of an allele pair match the observed sample reaction pattern See the discussion of Allele 1 Allele 2 columns below All samples that have been reviewed and accepted are assigned an alterna
133. ins a value Both cells are empty if no assignment can be made or if the assignments are ambiguous Version 1 1 69 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf 7 Comment sample specific comments you can select the contents of the comment cell by clicking at left side of the cell and dragging through all the text then copy and paste it to the Clipboard in the usual manner Analysis Date date of the allele assignment by LTI 9 Batch Analysis Comment a truncated version of batch specific comments Complete batch comments are printed out via the Lambda Reporter 10 Batch Name session ID assigned at the time of the Luminex run or updated upon import into LTT 11 Run Date date of the Luminex run Figure 6 1 Patient Typings Table 1 _ Patient ID 2 DOB 3 LastName 4 Comments i Jenkins9576 07 02 1974 Jenkins This Sya is half brother to older sibling from different mothers Anderson4210 13 01 1973 Anderson f p Klein3113 22 12 1986 O Consensus llele Consensus llele2 6 Comment 7 AnalysisDate Batch nalysis Comment BatchName A T1HGY A 33DWH No False Reactions 18 03 2005 4 new batch Augl4works CE E E EE 25072008 ERAN CE OC E A Rta EE DI E ELEVA ELE ESSE 2807208 Patient ID DOB LastName Comments Jesperson6 85 20 04 1967 Jesperson EE IC E EX A EC ELE CEI EA A You can modify the order of the columns of this table as discussed in Changing Batch Results Col
134. is al ways a positive number For A2 alleles i e with a 0 A1 score PP is the negative of the number of positives on the right side of the positives field in common with the sample reaction When the change in PP from the initial A2 allele PP score is greater than 2 a score of 0 is assigned 4 FN scores are For each allele the number of False Negatives plus 1 By definition the Sample Reaction is 0 5 TP scores are For each allele the number of True Positives The Sample Reaction is assigned a TP score of 101 which is larger than any possible number of True Posi tives No assay has more than 100 different types of beads This assures that the Sample Reaction always sub sorts as the first item in the 0 set of False Negatives 6 The alleles are then sorted and subsorted in this order A1 descending A2 descending PP ascending FN ascending TP descending To illustrate the sorting protocol Table B 6 reproduces part of a reaction grid with alleles scores for Al A2 PP FN and TP Deleted columns and rows are indicated by the tilde sign Version 1 1 127 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Table B 6 Allele Sorting in the Reaction Grid Allele 11 31 52 54 64 43 47 56 57 72 75 20 21 23 38 39 46 60 69 77 58 63 66 A1 A2 PP FN TP A 0302 KOK A RK XIX X x X x 5 0 10 2 11 A 0305 mee SG Gay Se 9p Fe OS USE rep oe h OK 5 0 11 1 12 A 0307 Ne Kara a E LG Se
135. isting of the new database the backup and the temporary copy of your original Thus if you run Update Database frequently the proliferation of backup and temporary files may consume a great deal of disk space 4 To avoid this and to protect your valuable data we recommend that you save the backup copies of your databases by burning them onto a CD or other storage medium before deleting them from your system 5 Before you copy or delete a database you must detach it from your MSDE SQL server instance select Start gt Programs gt One Lambda gt HLATools Configuration Suite gt Database gt Attach Detach from the Windows desktop 6 Select the Detach Database option Version 1 1 129 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf 7 Select the attached database from the pull down list and click Detach Figure B 12 Attach Detach Database Tool E HLATools Configuration Suite y o xj File gt 001883 SEB OLD ONELAMBDA HLA Update Comments Configuration Attach Database Connections Attach a database file as a database name Database Backup Restore Using the filename Local Create SS AA E Merge System Diagnostics User Management Detach Database Select a database on the server Detach How do I update an LTI database with a non default name The name for the LTI database is set by assigning a
136. itive Control Values panel and the Reaction Pattern Histogram Tabs at the bottom access the other principal views and return to the Batch Analysis View The views and subviews that address sample analysis tasks are shown schematically in Figure 5 7 A similar schematic for batch related views is shown in Figure 5 1 Figure 5 7 Assignments View and Subviews HLATools Batch Sample Main Panel Right Panel Explorer Bar Tree Five Tabbed Subviews Two Tabbed Subviews Matched Reaction Pairs Sample Assignment All Alleles Patient Information Allele Type Subtype Closest Reactions Bead Specificities Positive Reaction Pattern Histogram Controls Panel Return to Batch Analysis and Tabs to Four Sample Views Batch Analysis Assignments Bead Analysis Reaction Grid Raw Data Important Note In the Assignments view the vertical tabs to access the five main panel subviews may be initially hidden on small monitors or if the window is not maximized On large monitors an open field may appear between the five vertically tabbed subviews in the center and the two horizontally tabbed subviews on the upper right This extra space has been provided to accommodate small monitors 46 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Sample Results Analysis Assignments View with Matched Reaction Pairs and Close Reactions The Matched Reactions subview is one of a set of five vertically tabbed subviews accessible through t
137. ivided into three principal parts these introductory remarks software installation and networking instructions and usage instructions for the various HLATools software modules The manual also includes appendices dealing with topics in interface customization FAQs multi user installations a Glossary and an Index Page numbers in the on line version refer to page numbers in the printed version of the guide Chapter 2 Installation and Log On contains step by step instructions for installing the software on a single user standalone workstation Chapter 3 LABType Home provides a brief survey of the One Lambda product literature that forms the basis of much of the allele specific information that is contained in your HLATools database Chapter 4 Lambda Explorer describes how to access Luminex files and import them into the HLATools database Chapter 5 HLATools LABType Interactive describes how to use the LTI software application to analyze batch and individual patient or sample results and how to interpret allele assignments Chapter 6 Patient Explorer describes how to collate and review the results of multiple sample tests that have been carried out on a single patient Chapter 7 Reference Files and Their Maintenance describes the contents of several important data files which form part of the HLATools database and how to update them Chapter 8 HLATools Lambda Reporter provides an overview of the tools available to create
138. l In various LTI tables homozygous MRPs are indicated by an entry only in the Allele 1 cell The Luminex output files provides a number of different quantities that can be used as average readings for the beads e Mean the mean is the average calculated using all the data points for each bead For the dataset consisting of the five data points 3 5 6 7 14 the mean would be 35 5 or 7 e Median the median is the value of the data point which separates the upper half of the data set from the lower half If there are an even number of data points in the set the median is typically calculated as the mean of the two middle values For the data set cited above the median would be 6 Trimmed Mean the mean is calculated after a specified percentage of the high and low readings for each bead are thrown out to reduce the influence of outliers the de fault value for trimming in a Luminex output file is 5 each from top and bottom For the data set cited above trimming the top and bottom values would yield a trimmed mean of 18 3 or 6 Trimmed Median the median bead reading determined after trimming a given per centage off the ends of the data set For the data set cited above the trimmed median will be 6 The trimmed median and the median are generally identical The Merge Database function combines two LTI databases and in the process archives both individual databases If both databases contain the same batch the two in
139. l probably be Local Click Modify The information in Sections 1 and 2 will probably not need to be changed In Section 3 select the Attach a database file as a database name option to enable the two edit fields In the first field enter LTT without the quotation marks aA PP oY e Use the browse button to the right of the second field to locate the database file The pathname is typically Pro gram Files Microsoft SQL Server MSSQL ONELAMBDA Data LTI mdf Click Set to attach the database po 9 A user with administrator permissions can now run Update Database Remote LTI database Why can t connect to using Windows XP The Windows XP firewall on your computer may be preventing you from accessing the remote database You may need to open a port through your firewall In addition the local security policy on the remote computer may need to be changed from its default setting In Windows XP the default security setting forces all local users to access the LTI database as Guest It is necessary to change the security settings on the remote computer to allow other users on the network to authenticate as themselves rather than as Guest You must have administrative privileges to change the firewall port settings Open a port though the XP firewall How do 1 From the Windows XP desktop select Start gt Settings gt Control Panel gt Windows Firewall gt Exceptions If there are no entries
140. layed in the Reaction Pattern His togram Figure 5 8 only Bead the X symbol indicates the Allele 1 reaction pattern Bead the symbol indicates the Allele 2 reaction pattern Bead the diamond symbol indicates that the bead reaction is common to both Allele 1 and Allele 2 Bead the downward pointing arrowhead indicates that the cutoff has been increased with the result that the orig inal positive has become a negative Arrowheads point in the direction of the original value of the cutoff Tabs return to the Batch Results Table or access the other principal sample views e Bead Analysis this view is identical to the Batch Bead Analysis Rxn Grid a serogram like display of alleles and their associated bead reactions and false reactions See Reaction Grid p 58 e Raw Data a tabulation that includes the data shown in the tool tip 24 plus the probe recognition site for the bead and an abbreviated listing of bead specificities See Raw Data Table p 64 50 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 5 9 Assignments View with Two Close Reactions 6 DNA 8 87071 8 5001 Print Serological B50 Restore E Bw6 e Os Locked 11 Filter Unfitered y Comments Matched Reaction Pairs 2 3 Allelet Allele2 FalseReactions B OZAMYN gt EN B5001 4 eroro201 B 0723 B 5001 B 070203 B 0729 B 5001 B 070204 5 B 070202 B 5001 B 0715 B 0721 B
141. le reaction itself Figure B 5 Reaction Grid Default View Sample 5 Allele gt Alleles Print DRB1 01EW 7 DRB1 14WHG Pin lt gt Go Ae 341517 8 9 10 11 1214 15 16 17 18 19 20 21 2223 25 x x x x x ey tok Rxn DRB1 14WHG DRB1 1441 DRB1 1446 DRB1 01EW DRB1 010103 DRB1 1409 DRB1 1406 x xX KK KK OM e The y axis lists all the alleles in the assay Many of the entries in the Allele column are single unambiguous alleles which have wholly numerical designations Other entries designate groups of alleles with shared specificities and are designated with alphanumeric codes e When multiple alleles have a common reaction pattern the ambiguity is indicated using an NMDP or local ambi guity code For example in Figure B 6 the alleles DRB1 010101 DRB1 010102 DRB1 0107 DRB1 0107 DRB1 0108 and DRB1 0111 all share a common reaction pattern This group of alleles is designated in the Allele column by the NMDP code DRB1 01EW The member alleles designated by an ambiguity code are displayed in the table header space when you click anywhere in an allele s row e LTI compares the observed bead reactions with specificities represented by the alleles in the Allele column and marks the positive reactions with a red x 120 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure B 6 Reaction Grid Golden Alleles Sample 5 DRBWOIEW gt DAB1 010101 DRB1 010102 DRB1 010
142. ll newly entered values or changes into the database e Cancel discards all newly entered values or changes without saving them into the database The other fields are self explanatory and do not require elaboration Using Lambda Reporter Filters Selecting Samples The Lambda Reporter provides filters that allow you to specify very precisely which samples will be included in or excluded from your reports To illustrate these filters we will create a Patient Report that contains several samples 1 Select Patient Reports in the bottom panel of the Lambda Reporter sidebar to access a panel containing patient report search options Figure 8 4 Figure 8 4 Patient Reports Search Options Patient Reports Patient Report will s Patient Report Search Search Sections Paper Size Logo Inbox Archive Batch ID x Patient ID y Sample ID Catalog ID x Allele NMDP Specificity x QO Search 4 Uncheck All E Preview amp Export 2 Note that you can select Patient results from archived batches Archive as well as active batches Inbox In this example we will select both options 84 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 3 By default the Reporter matches up all patients from all batches Figure 8 4 We can use the Patient ID pull down list to narrow the search to a single patient The pull down list contains the names of all the patients in th
143. ls Configuration Suite 2 Place the CD ROM into your CD ROM drive 3 Many Windows computers are configured so that CDs auto run by default In the event that auto run has been dis abled on your system select Start gt Run gt Browse to locate your CD ROM drive 4 Double click on the CD ROM icon to launch the installation program If the Program Maintenance dialog box displays the following message you are reinstalling HLATools over an existing installation In that case proceed to Modifying Repairing or Uninstalling HLATools p 19 otherwise continue with the next step Figure 2 1 InstallShield Modify Message Welcome to the InstallShield Wizard for One Lambda HLATools The InstallShield R Wizard will install One Lambda HLATools on your computer To continue click Next 6 Once the installation begins you may see this message Figure 2 2 Microsoft NET Framework Installation InstallShield One Lambda HLATools uses the Microsoft R NET 1 1 Framework Would you like to install it now Yes No This occurs only if the software detects that Microsoft NET Framework has not been installed on your computer If Microsoft NET Framework is already installed this step of the install is skipped Tf the installation software cannot detect the Microsoft NET Framework software it will launch the InstallShield for Microsoft NET Framework Accept the license agreement click Install and wait for the installation to be completed 7
144. lse reaction removed from yielding an MRP See Closest Reactions Subview p 55 The False Reactions in Figure 5 8 involve only a single bead so they also appear in Figure 5 13 Closest Reac tions Subview e Specificity the allele specificities for each bead in the sample Mousing over a bead in the Reaction Pattern Histogram causes its specificities to appear in this subview See Specificity Subview p 56 DNA displays the allele assignments when at least one MRP has been found When more than one MRP has been identified the allele codes in the left and right hand DNA cells combine all the Allele and Allele2 assignments in the MRP table columns into a single code In Figure 5 8 both MRPs have the same allelel NMDP code so this code is shown in the left hand cell At the time of the analysis there was no NMDB code that would include both B 52AD and B 520101 so these are listed separately in two distinct MRPs In the right hand DNA cell combined B 52AD and B 520101 are represented by a local ambiguity code B 52XX2 Serological and Bw or Cw 1 Cw2 display serological equivalents and for B locus or C locus typing associated Bw4 Bw6 or Cw1 Cw2 allotypes of the HLA allele pairs shown in the All Alleles subview these fields remain blank if no serotype equivalent can be found or if the corresponding alleles are null alleles e g BBlank or CBlank If the B locus or C locus allelic subtype has been identified on the molecular level but not s
145. ment Subview Please see Figure 5 8 52 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf All Alleles Subview In contrast to the Matched Reaction Pair subview which employs NMPD or local ambiguity codes the All Alleles table displays all possible allele pairs down to the intron polymorphism level and lists them with their serological equivalents Figure 5 11 See Figure B 9 for a discussion of the method used to construct this table Figure 5 11 All Combinations of Both Allele Types DNA B 44AJVH 8 4801 Bie Serological Restore B44 B48 Bw Bu4 Bw6 JV Locked Filter Unfiltered Comments Recognized Allele Pairs Serological Equivalent Allele1 Allele2 Serol Sero2 Bw 1 amp B 4402010 B 4801 B44 B48 Bw4 Filter B 4402010 B 4801 B B48 But B 440202 BY4801 B48 But B 440203 B 4801 B44 B48 Bw4 B 4419N B 4801 BBlani 1 B48 Bw B 4423N B 4801 BBlank B48 Bw B 4427 B 4801 B48 B 4433 B 4801 B B48 B 4434 B 4801 B B45 No False Reactions 3 Matched Reaction Pairs B 444JVH amp B 4801 gt B 44020101 B 440201025 B 4419N B 4423N B 4427 B 4433 B 4434 amp B 4801 R 440203 amp R 48M1 gt R 440203 amp R 48M11 zi Previous Allele1 Allele2 lv Sample ID Sample 5 B 4402010 B 4801 Filter Date 4 14 2005 B 4402010 B 4801 Birth 5 12 1987 5 B 440202 B 4801 Race Cauc B 440203 B 4801 Patient Kleebold7324 Q ae a hd a E
146. move the program Sodify Change which program Features are installed This option displays the Custom Selection dialog in which you can change the way Features are installed Repair Repair installation errors in the program This option fixes missing or corrupt files shortcuts and registry entries C Remove S Remove One Lambda HLATools from your computer T InstallShield lt Back Cancel Modifying HLATools To modify HLATools software applications 1 Select the Modify option shown in Figure 2 5 2 Click Next to access the Custom Setup dialog Figure 2 6 e Installation options that will be offered in future releases can be accessed via the pull down menu e Ifyou select an option and the message in the Feature Description field declares that the feature requires no space on your hard drive this means that the feature is already installed 3 Click Next to proceed then Install then Finish to carry out the modification 20 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 2 6 Custom Setup Options ji One Lambda HLATools InstallShield Wizard Custom Setup Select the program features you want installed Click on an icon in the list below to change how a feature is installed HLATools Features R Hig Description This feature will be installed on local hard drive 58 This feature and all subfeatures will be installed on local ha
147. mple Data Control Values Tabs to Batch Views Batch Results Bead Analysis Control Values Version 1 1 35 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Batch Data Analysis You can access the LTI analysis module via the LabType Interactive icon in the Explorer Bar The batch sample tree will appear similar to that shown in Figure 5 2 The batch sample tree contains three top level folders You can transfer a batch from one folder to another by selecting the batch within the folder and using the right click menu options Inbox active batches csv session files that are currently loaded into the HLATools database and available for review Sample patient and batch data in active batches are included in reports To prevent information from a batch from being included in a report move the batch into the Archive folder Deleted inactive batches that will be permanently removed from the database the next time the Clean Database utility is run Archive inactive batches that you do not wish to discard Archived batches are not removed from the database by the Clean Database utility You can transfer Archived batches back into the Inbox at any time The colors applied to the batches and samples indicate the analysis and allele assignment status Black bold face sample contains a Matched Reaction Pair MRP but has not been reviewed and accepted Red bold face sample either has a false reaction or
148. mple in the selected batch The test date name of the tester csv filepath and batch comments are included in the header The report for each sample includes e Batch product lot information and locus type e Corrected accepted typing if any for each sample e Reaction assignment NIH 8 1 format Computer assignment combining all MRP designations into a single NMDP code pair if applicable e Generic allele descriptions with complete listing of types and subtypes for each MRP e Listing of close reactions Expansion of NMDP codes for computer allele assignments 4A Sample Raw Data Abbreviated Specificity Filtered by Batch This report contains raw data for the specified sample in a single batch e Batch product lot information and locus type e Draw test and analysis dates for the sample e Session ID and csv filepath for the batch 98 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf e Batch comments e Corrected accepted typing if any for the sample e Reaction assignment NIH 8 1 format is included in data tabulated table e Computer assignment combining all MRP designations into a single NMDP code pair if applicable e Generic allele descriptions with complete listing of types and subtypes for each MRP e Listing of close reactions Expansion of NMDP codes for computer allele assignments e Tabulated data for each bead in the sample Note in the table footer indicate whether a bead s positive cont
149. ms and the default values Just before finishing the installation the InstallShield Wizard creates an Upgrade LABType Database icon on your desktop You can launch the Upgrade LABType Database application in either of the following ways e Select the Launch LABType Import checkbox then Finish or Exit the installation by clicking Finish then launch the application using the icon 3 IfLABType is installed in the default CALABType directory you can proceed directly to the upgrade step If not a directory browse window will appear Browse to locate the directory containing the LABType executable labtypeN exe where N is the version number Figure B 15 132 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure B 15 Locating the LABType Classic Folder Browse For Folder 2 xi Browse for the LABType Classic that you wish to upgrade 314 Floppy 4 Local Disk C H E Documents and Settings H E labsern a LABType_new a LABType_old 3 LuminexFiles E MyBatches 3 olilat 3 Program Files O Seagull a SWSetup a temp tmp H E WINNT fa Cancel New Folder 4 H 5 laa labtype_advsetting mem 1KB MEM File Y labtypes exe 20 807 Application Ba lastesvdir mem 1KB MEM File LMTS INI 1 KB Configuration Settings CRUE 4 The installation program will check to see if the ONELAMBDA MSSQL server instance has the proper
150. n its positives are pulled to the left The rest of the grid is defined as the right or the Allele 2 remainder Two scoring cases are distinguished A No specification is made in the Second Allele entry field e The sample reaction is always scored 5 e Alleles that share more than 75 positives with the Second Allele are scored 5 e Alleles that share fewer than 75 positives with the Second Allele are scored 0 126 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf B A specification is made in the Second Allele entry field e The sample reaction is always scored 5 e Ifthe type or subtype designation first two and fours digits respectively of the allele matches that of the Second Allele and the allele shares more than 75 positives it is scored 5 e If the type or subtype designation first two and fours digits respectively of the allele does not match that of the Second Allele but the allele shares more than 75 positives it is scored 3 e Alleles that share fewer than 75 positives with the Second Allele are scored 0 3 PP scores can be assigned after the positive reactions have been clustered to the left of the grid The sample reaction is always scored as TPs in Sample Reaction TPs in First Allele This is always a neg ative number For alleles with a positive Al score PP is the number of true positives shared with the First Allele This
151. ng shared amino acid sequences A single antibody may react to many different HLA antigens if they share the same amino acid sequences Antigens with shared epitopes are called cross reactive groups CREGs A bead reaction missing from the sample observations An actual false negative is most often due to the sample reading being under the cutoff value When one compares the negative reading to other sample data and to QC data one may conclude that it is justified to adjust the cutoff value downward to allow the negative reading to be reclassified as positive A positive bead reaction that is not expected to be positive False positives are often the result of well contamination or an error during the preparation of the sample particularly during the PCR phase with the result that the sample reading is over the cutoff value When one compares the positive reading to other sample data and to QC data one may conclude that it is justified to adjust the cutoff value upward to allow the positive reading to be reclassified as negative After LTI searches for allele pairs with combined reaction patterns that yield exact Matched Reaction Pairs MRPs it searches for allele pairs whose combined reaction patterns differ by one bead reaction either positive or negative from yielding an MRP Such an allele pair may be referred to as a one away LTI records the allele pair and the bead ID of the bead involved in the false reaction LTI then searc
152. ng to LABScreen testing and negative control beads with no antigen attached e In Micro SSP a negative control reaction tube that detects the human beta globin in ternal control PCR amplification product this provides a background value since the beta globin gene amplification product is present in every well and is the most likely PCR contaminant A scoring scheme in which serological reactions are assigned values ranging from 0 no reaction to 8 very strong positive See Table Gl 2 for a tabulation of scores and their significance An ambiguity code currently consisting of from two to four alphabetic characters The characters in an NMDP code correspond uniquely to a concatenation of numbers used to designate allelic subtypes that share a common reaction pattern For example the NMDP code CKB corresponds to the numbers 01 03 07 Thus the ambiguity code B 45CKB is a shorthand designation for the allelic subtypes B 4501 B 4503 and B 4507 The assignment of NMDP codes is quasi random with new codes being created as longer or different numerical concatenations are required For example KBY gt 01 02 03 04 05 06 07 08 09 10 11 12 HDY gt 01 02 03 04 05 06 07 08 09 10 11 12 13 FYH gt 01 02 03 04 05 06 07 08 09 10 11 12 13 14 An example of four digit allelic subtypes including one null allele would be ASKK gt 0301 0303N 0304 0306 0307 0308 0309 A full listing of current NMDP codes can be found in the NMDP gt View which is
153. nguage differences that were not recognized by the installation utility There are several ways to identify which version and product level of MSDE is installed on your computer 1 From the desktop select Start gt Settings gt Control Panels gt Add Remove Programs 2 Locate the Microsoft SQL Server Desktop Engine ONELAMBDA entry 3 Click on the entry to expand it then click on the support information hyperlink The correspondence of version number and product level is as follows 114 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Table B 1 MDSE Version and Product Level Version Product Level 8 00 194 RTM 8 00 384 SP1 8 00 534 SP2 8 00 760 SP3 8 00 766 SP3a Alternatively select Start gt Run from the Windows desktop and execute the svrneten command to access the SQL Server Network Utility This requires administrative privileges Open the Network Libraries tabbed window The version numbers of the networking protocols are listed here Figure B 3 Figure B 3 Networking Protocol Versions Server network library Library file n File date Size Named Pipes SSNMPN7O DLL C PROGRA 8 0 534 12 18 2002 25 148 Bytes TCP IP SSNETLIB DLL C PROGRA 8 0 766 2 19 2003 82 492 Bytes NWLink IPX SPX SSNETLIB DLL C PROGRA 8 0 766 2 19 2003 82 492 Bytes If you do not have Service Pack 3 installed update the ONELAMBDA instance of MSDE as follows 1 Log in as administrator
154. no allele assignment has been made Figure 8 6 Samples to Report On 2 Sample s To Report On Include X Well Y SampleID Y Alelel Y Alel Y Comment Y Product Y M 2 B1 2436135 LowBead Count RSSO1B_006 Y 4 D1 2436137 LowBead Count RSSO1B_006 Version 1 1 85 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf 6 Further filters for searching on the values in each column are contained in a right click menu that can be accessed by clicking directly on the filter cone icon Figure 8 7 The right click menu lists each item in the column plus options for selecting all items All items with a value NonBlanks or no value Blanks Figure 8 7 Further Sample Table Column Search Filters 2 Sample s To Report On Include X Well Y Sample ID llelel VW gt Allele2 VW Comment gt Product Y Y 2 B1 All LowBeadCount RSS01B_006 vy 4 D1 ns Low Bead Count NonBlanks 2436135 2436137 7 The right click menu of Figure 8 7 also contains a Custom option that accesses a dialog with advanced filter crite ria Figure 8 8 With the Custom filter option you can create very specific searches for including or excluding samples In Figure 8 8 two conditions are applied to the search the Patient ID must be greater than Sample B4 and less than or equal to Sample B7 Note that a code representation of the selected search filter appears in the text field at the bottom of the dialog box
155. nt cutoff that defines a positive reaction when a bead cutoff has been modified from the original value sufficiently so that the bead reaction changes from positive to negative or negative to positive the back ground color for the Rxn and Data cells for that bead changes to light purple Table 5 3 Raw Data Table Background Colors Color Description Positive bead reaction Negative bead reaction or positive control bead Low bead count low positive control modified cutoff value has flipped reaction Version 1 1 65 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 5 26 Minimum Bead Count in Raw Data Table Raw Data Recognition Site 45 49 68 D Ste ews Taal Qi aa m1 Ha ad ada a PESAN ESE AG IAE ES ALE Bead counts that fall short of the threshold are indicated by a purple background in the table cell Figure 5 27 Figure 5 27 Low Bead Count in Raw Data Table Raw Data Recognition Site 93 W R 99 Spe perm at 30 EE ELEL Fs 1 fieses 69 2f o EE 19 24 Me 1 mennens 2 o E 19 14 les 8 62R as 68 150 33 A 4 e NC negative control which provides a measure of the background noise e PC positive control Class I products are assays for samples that contain two exons thus there is a positive con trol for each exon Since a given bead will be specific only for a site on a particular locus on one or the other exon different beads can hav
156. oad the users on the network in this case network1 into the display field 5 Select the name of the remote user who will be accessing this local database in this case Quimby Then press Add Quimby is now added to the list Figure C 5 140 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure C 5 Newly Added User with Access to Local Database HLA Tools Interactive Configuration Suite me ac File Analysis gt swordfish ONELAMBDA Comments II sa Configuration BUILTIN Administrators Connections swordfish workshop1 b swordfish tissuelab Database swordfish Homer Attach Detach network1 Quimby Backup Restore Clean Update Merge Diagnostics List Users User Management Add SQL User Add Windows User Group Delete User 6 The LTI database on swordfish can now be accessed by two default local administrators sa and BUILTIN Administrators three local users workshop1 tissuelab and Homer and one remote user Quimby Now that Quimby has been added to the authorized users list on Homer s computer swordfish log into Quimby s computer skipjack and launch the Connection utility by selecting Start gt Programs gt One Lambda gt HLATools Configuration Suite gt Connections Later we will see how to do this remotely Version 1 1 141 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure C 6 HLATools Configuration Suite Connections Tool E HLA Tools In
157. oding code page 65001 e UTF 7 encodes all Unicode characters using 7 bit encoding code page 65000 110 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf AppendxB HLATools FAQ This appendix addresses a number of issues that may arise when you are setting up or using HLATools LABType Interactive Frequently Asked Questions How can tell if HLATools is using the most recent LABType products 1 Check the product listing displayed in the HLATools Product Information table which you can access by launching the HLA Tools Configuration Suite and then selecting HLA Update gt View 2 In your internet browser access the One Lambda website http download onelambda com pub tray_info Windows HLATools Labtype_Interactive and compare the product listing The most recent LABType products will appear on the download site and they will also appear in the HLATools Product Information table Figure 7 2 For more on product revision numbers see Products gt Update p 75 Why can t I find the Admin Diagnostics tool In the Version 1 0 release of HLATools many of utility functions were carried out by standalone applications such as the Admin Diagnostics Tool and the Quick Connect Tool These and a number of other utilities have been combined into the HLATools Configuration Suite which is accessible by selecting Start gt Programs gt One Lambda gt HLATools gt Configure from the Windows desktop For exampl
158. of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 21 22 23 24 25 26 27 28 29 made to Close Reactions or reactions with multiple false reactions to obtain an MRP For color assignments in the histogram see Table 5 1 Bead Analysis Color Codes Reaction Pattern Histogram Symbology x Positive reactions in Allele1 Positive reactions in Allele2 Positive reactions present in both alleles Cutoff value A Cutoff adjusted downward forcing a negative reaction to become positive points toward original setting Cutoff adjusted upward forcing a positive reaction to become negative points toward original setting Selecting a cell in the Allele 1 or Allele 2 columns in the Matched subview causes the Reaction Pattern Histogram to display the symbols for the beads in the sample reaction pattern that belong to that allele Figure 5 10 It also causes the allele assignment in the header to display the full code definition 4 Clicking Restore 9 undoes any user initiated changes to the bead cutoff values and restores the original assignments Y axis normalized bead reaction values percent of cutoff value see Eq Gl 1 X axis bead ID numbers control bead s are not included Cutoff adjustment adjust by dragging the bead symbol vertically the current adjustment appears in a tooltip Figure 5 9 only Tool tip mousing over a bead displays a tool tip containing the information disp
159. of the alleles have the first two digits in common The same method is applicable when the first two digits are different For the sake of simplicity only the first four digits are considered in this sorting routine Table B 4 Grouping Alleles for the MRP Tabulation Allele 1 0101 0101 0101 0101 0101 0101 0102 0104 0106 D O 0 1 Dn FW NY Original Pairing Allele 2 0101 0102 0103 0104 0105 0106 0103 0105 0106 Simplified Final Pairing Allele 1 0101 0103 0105 0106 Allele 2 0101 0102 0103 0104 0105 0106 Three rules are observed 1 Trivial duplicate entries in a column are avoided Thus in rows 2 through 6 0101 is not repeated in Allele 1 but 0102 0103 etc are displayed in Allele 2 because each instance is a first occurrence 2 The entries for the columns are swapped if one or the other column already contains the entry Thus in row 7 Allele 2 already contains an instance of 0103 forcing the entries to be swapped 0103 is now grouped under Allele 1 while 0102 is deleted under Allele 2 because there is already an instance of 0102 in that column The same holds true for the entries in r 3 A homozygous allele is represented in both columns row 9 ow 8 122 of 172 OLI UD 0157 062006 6 6 06 HLATools_IVDManual_v1 1_Rev 0 pdf In the MRP tabulation and elsewhere high resolution alleles are designated with four digits the first pair designates the
160. ols in a Client Server Network to come 151 HLA Tools Security Modes mcocicocerrrcrinrrrra ra a anaes 151 Sharing Luminex Files on a NetwOrk oooooooococoororororrr nee 152 COssary Entries sacrer isra ee nE RE RENER RARE ARALAR 157 A260 A280 Rill oare co td co 64 E es ed ate eG E E ile ae 157 Dj ckne taceeeeatedeeakacaraear ase bacedk ait abawacacuhasararks 157 Allele Name Extensions ci sees poe a eae EAA Shae SIRS 157 Amino Acid Codes ii eaed bo bae eaiae de aaa 157 nn anaa a e aea a e Ei 158 Class LMC oro Rs ia 158 Class ILMHC cr CSR eee ria 158 Cell d th detectin osease sesa ys dsp 159 Clean Database sir bees kenaka Ad A A Ad ca 160 Close R action 615 059 ai Seed eRe wd PS AS Be 160 Code definition Sa 160 Complement errar DA E E 160 COLOCADA a ERA ARE 160 CREG oiga rai 160 EDTA A o nn de E E 161 A chara bawaackbe ace aces 161 Epitope lt ise sede eer e o2 ease RESO RENERE eee Redes ee Eos ee ees 161 False 0 A oud tem ai anei eai e E A ae n EALE aa 161 False POSTE cocotero drena 161 Palse R action caw noes EVES oo URE ES oe Pee as eee 161 Gentle Code A HEE DERE DG ERED 162 Immunomagnetic beads 2 44152 ose esseeeued ibe ewegieeseeu se eeeees 162 DPI NAUK AAA ee oe Gee A eee A ee ale Ye aaa e eke ee ak GER ee es 162 Local Code o c2dse Seeded eS tripea eer obs ehh ee etre El eae 162 Eos Piles df e oe ae wate peace ew a9 Buea oe 162 Matched Reaction Pat ta dai 163 Meal anc Median iS Li AO AS RA 163
161. ols_IVDManual_v1 1_ Rev 0 pdf Listing of close reactions Expansion of NMDP codes for computer allele assignments 3B Batch Raw Data Abbreviated Specificity Report No patient data Batch product lot information and locus type Draw test and analysis dates for each sample csv filepath Batch comments Corrected accepted typing if any for each sample Reaction assignment NIH 8 1 single row format is included in raw data table Computer assignment combining all MRP designations into a single NMDP code pair if applicable Generic allele descriptions with complete listing of types and subtypes for each MRP Listing of close reactions Expansion of NMDP codes for computer allele assignments Tabulated data for each bead in the sample including e Bead number e Reaction status 1 or 8 e Recognition site with amino acid mutation e Raw data and normalized value for bead reaction e Cutoff percent e Values for negative and positive control s e Bead count e Abbreviated allele specificities for each bead 3D Batch Report Sorted by Patient This report omits any samples not associated with a patient ID All associated samples are listed under a patient specific data header Draw dates for individual samples are not included Patient data name gender etc and patient specific comments Batch product lot information and locus type Draw test and analysis dates for each sample csv filepath Batch comments 96 of 1
162. ols_IVDManual_v1 1_Rev 0 pdf 10 Allele 1 Allele 2 an exhaustive listing of allele assignments for the loci on the paternal maternal chromosomes Selecting a cell in either column causes the Reaction Pattern Histogram 20 to highlight the part of the sample reaction pattern that belongs to that allele Figure 5 10 False Reactions this column contains the IDs of the bead s that if their reaction state were flipped to the oppo site negative to positive or positive to negative would allow the alleles listed in the Allele 1 Allele 2 columns to form an MRP The False Reactions in Figure 5 9 each involve only a single bead so they are considered Close Reactions Code allele assignment in the selected cell in the Allele 1 Allele 2 columns when multiple allele share the same reaction pattern this is indicated by an NMPD or local ambiguity code Allele Listing the definitions of the code 3 are displayed in this area Tabs access four subviews e All Alleles a listing of recognized allele pairs and their serological equivalents See All Alleles Subview p 53 e Type Subtype a listing of all the allelic types and subtypes contained in recognized allele pairs See Type Subtype Subview p 54 and Type Subtype Allele Assignments How are made p 125 e Closest Reactions a tabulation of all Close Reactions for the current sample i e reaction pattern combina tions that are only one fa
163. olumns of beads with observed positive reactions are indicated by a light blue background e Aredxona light blue background indicates a true positive reaction e A black x on a white background indicates a false negative reaction for example in Figure 5 16 if a positive reaction for bead 82 had been observed the sample reaction pattern would also include bead 82 and allele DRB1 1109 would be part of the assignment e A clear cell in the Rxn row indicates a true negative reaction a clear cell in a column with a light blue back ground indicates a possible false positive reaction e The top row lists the probes by number The numbers of probes that were not used during data acquisition are excluded for the sake of compactness in this example Table 5 2 Reaction Grid Background Colors Reaction Color Description Positive Positive bead reaction if cell contains red x False positive if cell is empty Observed Sample reaction pattern Reaction Pattern Type color for Golden allele No Reaction No reaction False negative if cell contains black x Golden Allele Allele reaction pattern is subset of sample reaction pattern Excluded Bead Bead globally excluded from analysis The elements and controls in the Reaction Grid are enumerated below 1 Sample ID as distinguished from a Patient ID 2 Allele1 Allele2 suggested allele assignments if no assignment is possible the allele designations are left bl
164. om the cytoplasm of the cell These antigens are typically fragments of viral proteins or tumor proteins from within the cell itself Class I MHC molecules are made up of two chains a heavy chain transmembrane polypeptide and a light chain beta 2 microglobulin that does not penetrate the cell membrane Class I OLI products employ two positive control probes bead 13 for exon 2 and bead 32 for exon 3 Class II MHC Class II MHC molecules usually display peptides derived from extracellular proteins that are internalized into phagocytic or endocytic vesicles These extracellular proteins are typically fragments of bacteria or viruses that have been engulfed and processed or broken down by a so called professional antigen presenting cell such as a macrophage 156 of 172 HLATools User Manual 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf or dendritic cell Class II OLI products employ a single positive control probe bead 34 for exon 2 Figure Gl 1 Class and Class II MHC Molecules Antigen Antigen Y al pl a2 p2 a a B Class MHC Molecule Class II MHC Molecule Antigen Presentation to Antigen Presentation to CD8 T cells CD4 T cells Cell death detection In serological screening cell death is detected by the entry of a stain into dead cells A commonly used staining medium is a mixture of acridine orange AO and ethidium bromide EB in a solution of bovine hemaglobin Acridine orange a green dye stains li
165. options 149 modifying policies 114 Serological equivalents B 77 BBlank 77 Server Network Settings 116 running svrnetcn exe 135 SQL server showing current instance 7 SSO Sequence Specific Oligonucleotide 5 SSP Sequence Specific Primer 5 Substrate in ELISA tests 165 Superparamagnetism 160 T Tables see also Figures Allele sorting in the Reaction Grid 128 Amino acid codes 156 Amino acid nucleotide triplets 160 bead profile color coding 40 creating the Type Subtype table 125 grouping MRP alleles 122 HLA allele nomenclature 116 examples 117 MSDE versions and product levels 115 NIH scores and cell death rates 157 Nucleotide codes 163 raw data table color coding 63 reaction grid color coding 57 TCP Transmission Control Protocol 112 TCP IP 116 136 Trimmed Count default setting 158 Trimmed Mean default setting 161 Trimmed Median 161 U UDP User Datagram Protocol 112 113 Unicode 165 Update Database database does not exist 112 170 of 172 OLI UD 0157 062006 HLATools User Manual Index 6 6 06 HLATools_IVDManual_v1 1_Rev 0 pdf execution denied 112 V UTF Unicode Transformation Format 109 Visible Utilities appearance priority in Product Select 72 73 see HLATools Configuration Suite w Wireless network installation 135 Version 1 1 HLATools User Manual Index 171 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf 17
166. ove the sample reaction pattern in the Rxn row by double clicking anywhere in its row Its final position is determined by its similarity to the observed reaction pattern Grouping alleles together in this way facilitates comparison with the sample reaction making it easier to see which combinations of alleles can form a Matched Reaction Pair Examples of pairs of alleles shifted for this reason were seen above in Figure 5 17 and Figure 5 18 Restoring Allele Order You can restore the allele listing to its original order with the golden alleles grouped at the top by double clicking on the Allele column header The other alleles are displayed in descending order in terms of their closeness of match Figure 5 22 62 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 5 22 Restoring Allele Sort Order PAE Rxn x x x x x E Os x DPB1 02CXD x DPB1 5201 x DPB1 1301 DPB1 2901 DPB1 7001 DPB1 8801 DPB1260102 DPB1 1401 DPB1 5001 DPB1 5601 x x x x x x KX KK KK KK OK OK Sorting by Bead Usage You can also sort the Reaction Grid to see how many times a given bead was involved in allele reactions by clicking on the bead number in first row Figure 5 19 shows that bead 51 was involved in eight allele reaction patterns Clicking on the bead number or anywhere in that bead s column also displays the allele specificities for the bead in the header panel Sorting by Allele Pattern Matches T
167. pdating process see the FAQ Why is the LTI database directory growing excessively Figure B 13 Setting Database Name in ALL bat lolx File Edit Format Help echo off SET SERVER local oneLambda SET TABLE HUGO SET LOGIN E REM SET LOGIN Usa Psecret REM SET SERVER 192 168 1 200 oneLambda net start mssqlfonelambda osql 5 XSERVER LOGIN q EXITCDROP DATABASE TABLE net stop mssql onelambda Del XProgramFilesiimicrosoft SOL Server MSSOLSONELAMBDA Dat a TABLEX net start mssqlfonelambda osql 5 XSERVER LOGIN q EXITCCREATE DATABASE TABLE Figure B 14 Setting Database Name in updateDB bat fj updateDB bat Notepad loj xj File Edit Format Help echo off Set server C local oneLambda Set Service MSSOLSONELAMBDA Set Login E Set Table HUGO setmydt gt templtiws call setmydate bat How do install MSDE to a non default folder You must have administrative permissions to perform this installation To Install MSDE to a Directory other than Default 1 Goto http msdn microsoft com sql 200 downloads default aspx 2 Click the MSDE 2000 Release A link 3 Select the language version you would like to download and click Change 4 Scroll down the page and click Download for the lt Ing gt _MSDE2000a exe file Version 1 1 131 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf where lt Ing gt is the 3 letter abbreviation for the language you hav
168. ple Raw Data Abbreviated Specificity Filtered by Batch 98 4B Combined Sample Analysis Data Report oo oooomomomo o 99 4C Sample Raw Data Abbreviated Specificity Report 100 4D Sample Abbreviated Specificity Rep0rt oooooooooooooo o 100 4E Sample Complete Specificity Report 0 0 00 00005 100 99A Sample Report with Chart Filtered by Batch 100 Summary REPO sexista yd eve ea nee dra ii 101 1A ABDR by Batch Sorted by Well Position o oooooooo 101 TC Bateh Results ens a casa marmane a iae aa a e S 101 5B Allele Query Report 2 44 4 hes4 cheeses bevaaeeeababaken ds 101 6A Serological Equivalent Table Report o ooooooooomooo o 102 Version 1 1 HLATools User Manual 3 of 172 OLI UD 0127 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf 6B NMDP Allele Code Data o o o o 102 6C Batch Data File Summary Report 0000055 102 6E Catalog Information 0 0 eee eee ee eens 102 Special REPOS s o2 lt oSecus oe eGa DAR A IAEA EEG 103 SA Allele Group Frequencies 02 cece ete e eens 103 6D Database Table Information 0 0 0 eee ee eee es 103 99B ABDR by Batch Sorted by Well Position 103 Appendix A LTI Interface Customization M difying Tables cosida dass Seas 107 Changing Sort Order or Grouping
169. ports If a Database Login dialog appears when you try to generate long reports such as the Patient Summary Report by Batch it means that the TCP IP and Named Pipes protocols have not been enabled in your Server Network Settings You can ignore the request for login However the time required to print out the requested report may be inconve niently long The steps required to enable these protocols so that all reports are generated and printed quickly are described in detail in Peer to Peer Installation p 135 Windows Authentication Mode Why can t I set up If your computer uses the Windows 2000 operating system and you have chosen Windows Authentication as your security mode as described in HLATools Security Modes p 149 you may find that you are unable to log in to LTI This is due to the fact that Version 8 0 of the Microsoft SQL Server Desktop Engine MSDE does not support the Windows authentication mode on systems running Windows 2000 This problem will be rectified with an upcoming release of the MSDE software Alleles How are they designated The HLA locus A B C DR DP or DQ is designated first followed by an asterisk The allele type and subtype are designated by four digits The first two digits designate the serological equivalent of the antigen The second pair of digits designates the specific allele Additional digits may be added that indicate a separate higher resolution subtype that is produced by a synon
170. r in sample Rxn the reaction status of a bead is indicated by 8 positive or 1 negative A positive bead reading is also indi cated by a light red cell color For example bead 18 in Figure 5 25 has a Data reading of 192 which is above the 56 cutoff for that bead The cell color for positive control beads is always white to make them easily distinguish able from other beads with positive reactions e Recognition site these entries in Figure 5 25 contain the start and end points of the sequences that are recognized by the probes Also included are any amino acid polymorphisms that render the probe more specific For details on interpreting recognition site nomenclature see the Recognition Site glossary topic Sorting the table by recognition site provides an insight into the remarkable specificity of the probes Figure 5 25 Raw Data Sorted by Recognition Site Bead Axn Recognition Site CA E Cutoff Count PC 2 1 60 G 65 23 138 1729 MENEN EI IL E E II pajas a e ai 173 Ea eras ed e 0 17 Beads 18 and 28 differ only in that bead 18 specifies the substitution of a single amino acid N or asparagine at the 63rd position in the sequence In all other respects the bead sequences are identical e Raw the mean trimmed mean median or trimmed median value of the fluorescence readings of the bead before normalization e Data the normalized value of the reading this value is a percentage e Cutoff perce
171. rd drive This feature requires OKB on your hard drive Installshield Help lt Back Cancel Repairing HLATools If you encounter a runtime error message similar to that shown in Figure 2 7 you can run the Repair utility without completely reinstalling the program Figure 2 7 Runtime Error Message LTI exe Common Language Runtime Debugging Services A Application has generated an exception that could not be handled Process id 0x81c 2076 Thread id 0xde8 3560 Click OK to terminate the application Click CANCEL to debug the application cme Note that repairing the HLATools software does not affect your Luminex data files or database files 1 Select the Repair option shown in Figure 2 5 2 Click Next then Install The repair takes several minutes 3 Click Finish to exit Version 1 1 21 of 172 OLI UD 0157 062006 HLATools_ VDManual_v1 1_Rev 0 pdf Uninstalling HLATools You can uninstall HLATools either by selecting Start gt Settings gt Control Panel gt Add or Remove Programs from the Windows desktop or you can use the HLATools Uninstall utility 1 Select the Uninstall option shown in Figure 2 5 2 Click Next then Remove The uninstall takes several minutes and does not affect your Luminex data files or data base files 3 If you do not plan to use HLATools further on the computer from which you have uninstalled the applications you may wish to remove the Luminex data file
172. rk Utility 136 SQL user creating new 148 Synopsis of Batch Analysis 33 Synopsis of Sample Analysis 44 Type Subtype Tables 52 126 Updating LABType product information 73 User Management Tool 139 Windows network domain users 140 Windows XP changing security policy 114 modifying firewall settings 113 Firewall making a port through 112 G Genetic code 160 Golden allele 58 60 121 Version 1 1 OLI UD 0157 062006 H HLA allele nomenclature 116 HLATools installing new version 20 LABType Home 5 LABType Interactive LTT described 5 Lambda Explorer 5 Lambda Reporter 6 Patient Explorer 6 HLATools Configuration Suite Attach Detach Database 7 Backup Database 7 Clean Database 7 158 Comments Editor 6 complete uninstall of HLATools 20 Connections Tool 7 HLA Update 6 System Diagnostics 7 Update Database 165 User Management 8 l Immunomagnetic beads 160 Installation updated versions of HLATools 20 Internet Explorer security options 10 J JPN rank 160 L Lab Information Interface 81 LABType Classic importing analysis results into HLATools 132 Local code 160 Low expression alleles 163 LTI database showing currently attached 7 Luminex files C LuminexFiles folder 16 150 C MyBatches folder 150 setting sharing permissions 150 M Mean and Median defined 161 MIC 161 Microsoft NET Framework 13 Microsoft SQL Server service stopping 20 MRP mat
173. rk so that the user can import csv files which contain data to be analyzed store the results of analysis and ultimately create analysis reports When the combined reaction patterns of a pair of alleles are only one false positive or one false negative removed from yielding a Matched Reaction Pair MRP this is termed a Close Reaction An example is shown in Figure Gl 2 Here the combined reaction patterns of alleles DPB1 1401 and DPB1 1301 would be consistent with the positives in the sample reaction pattern Rxn if the sample had also exhibited a positive reaction of bead 12 Figure Gl 2 Close Reaction Allele 1 2 3 4 5 6 7 8 a 10 11 12 113 14 15 16 17 18 19 20 22 23 24 35 36 37 43 PP Alleles DPB1 1401 0 DPB1 1401 DPB1 1301 0 DPB1 1301 The NMDP code definition is the expansion of the code to show all the component alleles For example the definition of A 33DWH is A 3301 A 3303 3304 3305 3306 Complement A group of proteins in the serum that is activated by the antibody antigen complex on a cell surface The result may be the destruction of the antigen or lysis of the presenting cell Count Type The Luminex output files contains two different count data points Trimmed Count the count for each bead is reduced by a specified percentage the default value is 10 e Count the full count of each type of bead drawn from a well this is the default option used during the LTT analysis CREG Cross REactive Group
174. rol or count are below the minima specified in the HLATools Product Information table e Bead number e Reaction status 1 or 8 e Recognition site with amino acid mutation e Raw data and normalized value for bead reaction e Cutoff percent e Values for negative and positive control s e Bead count e Abbreviated allele specificities for each bead 4B Combined Sample Analysis Data Report This report contains the test results for all samples with the specified sample ID in all active batches i e batches in the LTI Inbox folder e Patient information for associated sample Only one patient can be associated with a given sample e Batch product lot information and locus type for each batch e Draw test and analysis dates for each sample e Session ID and csv filepath for each batch e Batch comments for each batch e Corrected accepted typing if any for each sample e Reaction assignment NIH 8 1 format is included in data tabulated table e Computer assignment combining all MRP designations into a single NMDP code pair if applicable e Generic allele descriptions with complete listing of types and subtypes for each MRP e Listing of close reactions e Expansion of NMDP codes for computer allele assignments Version 1 1 99 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 4C Sample Raw Data Abbreviated Specificity Report This report contains the same information as report 4A However whereas 4A contains
175. s software suite This manual supersedes the introductory topics that were treated in Getting Started with One Lambda HLATools and contains advanced topics and reference material that goes beyond the scope of that document All One Lambda software products are designed to assist personnel experienced in HLA analysis by suggesting typing results However any clinical or diagnostic results must be carefully reviewed by a person qualified in HLA typing to assure correctness The software may be used to aid in suggesting results but should not be used as the sole method for determining reportable results The software is meant as a laboratory aid not as a source of definitive results Overview One Lambda HLATooIls is a suite of software applications that interpret the results obtained by One Lambda s LabType SSO HLA typing tests The software suite contains five major modules and a number of utilities HLATools Modules The application suite comprises the following modules LABType Home a repository of One Lambda product lot specific information in PDF format for LABType SSO and Micro SSP product lines Lambda Explorer a utility for importing files produced by the Luminex LABScan lt SuperScript gt 100 bench top flow analyzer into the LabType Interactive LTT database LabType Interactive a software application that contains tools for assessing the quality of imported data reviewing preliminary analysis results adjusting c
176. s 2000 double click on the MSSQLSONELAMBDA service to access the MSSQLSONELAMBDA properties dialog and select Stop gt OK On Windows XP select MSSQLSONELAMBDA from the list of services and use the Stop option in the left panel or double click on the MSSQLSONELAMBDA service to access the MSSQLSONELAMBDA properties dialog and select Stop gt OK When you reinstall HLATools the MSSQLSONELAMBDA service will also be reinstalled and started automatically 4 Rename C Program Files Microsoft SQL Server MSSQL ONELAMBDA to old MSSQL ONELAMBDA 5 Delete the Start Programs One Lambda directory from the Start menu 6 Delete the entire folder C Program Files One Lambda You may now install the new version of HLATools 22 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Updating HLATools When you install a new version of HLATools you might be concerned that all your existing analysis results are preserved To address this concern HLATools automatically backs up your database when updating HLATools Follow the steps shown in Figure 2 8 to determine whether you should run Update Database Clean Database or neither Figure 2 8 Updating HLATools 1 Uninstall old HLATools 2 Install new HLATools 3 Update or Clean DB 4a Launch Update DB 4b Launch Clean DB 1 The name of the currently attached database is shown in the HLA Tools Configuration Suite Database panel Figure 2 9
177. s LabType SSO HLA typing tests The central module in the suite is the LabType Interactive LTI analysis tool The bulk of this document is devoted to explaining the use and features of LTI With LTI and the other software modules in this suite you can accomplish the tasks listed below Import data from the LABScan 100 bench top flow analyzer Importing Luminex csv Files p 31 Review preliminary results in tabular format Batch Data Analysis p 36 Assess the quality of the imported data by comparing it to demographic reference data Bead Analysis View p 42 Adjust cutoff values globally and locally as warranted Bead Analysis View p 42 and Assignments View with Matched Reaction Pairs and Close Reactions p 47 Accept suggested allele assignments Assignments View with Matched Reaction Pairs and Close Reactions p 47 Review identified Close Reactions cases where reversing a single bead reaction making a negative positive or vice versa would yield a Matched Reaction Pair and an allele assignment Closest Reactions Subview p 55 Review types and subtypes of assigned alleles as well as ambiguous cases where the beads cannot determine with precision which specific alleles are present and allele assignments cannot be made Type Subtype Subview p 54 Review allele specificities for any given bead in the sample Specificity Subview p 56 Identify and assign serological equivalents to the assigned allele All Alleles Subv
178. s from the C Luminex files folder and the HLATools database files from C Program Files Microsoft SQL Server MSSQL ONELAMBDA Complete Uninstall of HLATools In rare instances you may encounter problems running HLATools after installing a new version These problems may be the result of the incomplete removal of old program components and registry data during the uninstall If this occurs we recommend that you perform a complete manual uninstall of the old version and its supporting files and folders To do a complete uninstall 1 Uninstall all existing instances of HLATools the HLATools Configuration Suite and MDSE using the Windows Add or Remove Programs utility accessible by selecting Start gt Settings gt Control Panel gt Add and Remove Programs 2 Delete the HLATools folder in C Documents and Settings All Users Application Data OneLambda In many sys tem installations the Application Data folder is hidden If you cannot find it in the All Users folder you can access it by navigating to All Users and then entering Application Data in the Address field Alternatively select Start gt Settings gt Control Panel gt Folder Options gt View gt Hidden Files and Folders and choose the Show Hidden Files and Folders option 3 On both Windows 2000 and Windows XP stop the Microsoft SQL Server service by selecting Start gt Settings gt Control Panel gt Administrative Tools gt Services gt Services Local then e On Window
179. s themselves Consider the recognition site entry for bead 1 in Figure Gl 3 E de 14 probe MRYFFTSVSR reference sequence This entry indicates that the probe is specific for the nucleic acid sequence that encodes the amino acids in the pertinent locus at positions 5 through 14 with a substitution of tyrosine Y for phenylalanine F at the 9th position See Table Gl 1above for a listing of amino acid codes The reference sequence alignments for the various loci can be found at the Anthony Nolan Research Institute website under the category of HLA Class I and II Sequence Alignments See http www anthonynolan org uk HIG data html 164 of 172 OLI UD 0157 062006 HLATools User Manual 6 6 06 HLATools_IVDManual_v1 1_Rev 0 pdf RFLP Secondary antibody SSO Substrate Unicode Update Database Version 1 1 Figure Gl 3 Raw Data Table Recognition Sites i di Recognition Site nse Cutoff Count aa BE 60 G 65 23 292 E 8570101 EEE CI E a aaa 18200075 MEN ESC E A E E E Gh pasa I 002 erro BESNILA Ce a E a eee BC ap Restriction Fragment Length Polymorphism the first molecular typing method applied to the Class II MHC in 1982 RFLP analysis is based on restriction endonuclease cleavage at polymorphic restriction sites These sites specific for each enzyme endonu clease used are located in both coding and non coding regions of the genes The strong linkage disequilibrium between
180. sample 100 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Summary Reports 1A ABDR by Batch Sorted by Well Position This report returns analysis exports for A B Cw DQ and DR loci in Excel spreadsheet format Because of the width of the spreadsheet the preview will only display a small part of the report You must open the exported file in Excel to see all the information The report includes e Sample ID e Patient Name e Draw test and analysis dates for each sample e Catalog Lot number A B Cw DQ and DR assignments using NMDP code and numeric expansions of NMDP code e Accepted Not Accepted status 1C Batch Results This report contains all of the allele assignments and numeric expansions for the samples in the specified batch Infor mation includes e Session ID e Product Lot for batch e Sample IDs e Well locations e Allele assignments and numeric expansions for each sample e Associated patient and kinship information e Accepted Not Accepted status Number of Close Reactions Number of MRPs 5B Allele Query Report This report returns all of the samples in which the specified allele has been detected Only accepted samples are included in the report The report includes for each sample e Catalog Lot number e Draw test and analysis dates e Patient data for the sample e Reaction assignment e Corrected typing Version 1 1 101 of 172 OLI UD 0157 062006 HL
181. serotype for Allele 1 and Allele 2 CR count of Close Reactions a close reaction is one that is just one bead reaction status change away from yielding an MRP See Closest Reactions Subview p 55 MRP count of Matched Reaction Pairs Comment II user supplied stock comment applied using the Comments pull down menu Figure 5 8 12 Tech1 Tech2 usernames of preliminary and confirming reviewers Version 1 1 39 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf e Datel Date2 time stamps for preliminary and confirming reviews e NMDP1 NMDP2 definitions of NMDP or local codes used when Allele 1 and Allele 2 have been re viewed and accepted e A1 4Digit A2 4Digit type subtype assignments for Allele 1 and Allele 2 the assignment reflects the most frequently observed subtype 12 Bead Analysis accesses the Bead Analysis View which compares sample batch bead profiles with QC refer ence bead profiles For details see Bead Analysis View p 42 13 Control Values accesses the Control Values view which displays values for the control beads Control Values View Positive Controls and the bead counts in a sample Control Values View Minimum Bead Count 40 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure 5 4 Bead Analysis View I23_104B_L6 Bead 59 59 E orri Exclude Bead o il from Analysis o 30 G Globally Changed Cuto
182. specific polymorphic restriction sites and coding sequence variations allows RFLP analysis to be useful in HLA Class II DNA typing A good corre lation exists between DNA RFLP analysis and phenotypic typing for Class II specificities Early forms of RFLP analysis using radio tagged probes have been largely replaced by PCR based analytical methods In ELISA assays the detection antibody that binds to the serum antigen which has already bound to the purified capture antibody attached to the bottom of the well The secondary antibody in turn becomes bound to the colorimetric substrate The strength of the signal from the substrate can be used to make a quantitative determination of the amount of antibody in the sample Sequence Specific Oligonucleotide SSO typing involves PCR amplification of a chosen sequence using primers flanking the sequence The amplified DNA is immobilized on a membrane and hybridized with selected oligonucleotide probes that are labelled with radioactive S or 7P by ELISA or by chemical markers In ELISA assays the substrate is the chromogenic chemical species that produces a signal when it binds to the secondary antibody One Lambda ELISA assays typically use either pNPP para nitrophenyl phosphate or BCIP 5 bromo 4 chloro 3 indolyl phosphate as a substrate A standard for representing characters as integers Unlike ASCII which uses 7 bits for each character Unicode uses 16 bits which means that it
183. stances are kept separate since each batch in the merged result is assigned a different internal identifi cation number If you merge databases on a regular basis Lambda recommends that you perform the merge whenever the from databases contains a thousand or more samples MHC Class I Chain related markers appear to play a role in graft rejection MIC proteins are single chain polymorphic proteins Currently 6 2006 there are 51 variants and approximately 60 recognized alleles Since MIC gene products are expressed on endothelial cells but not on lymphocytes conventional cross match procedures do not detect anti MIC antibody HLA Cw4 and Cw6 belong to one allotype group of HLA C proteins characterized by the presence of K80 lysine at the 80th residue whereas Cw3 and Cw7 belong to the other group that has N80 asparagine at the 80th residue A control intended to provide a reference value for no reaction whatsoever Negative control values are typically used to confirm that the data collection methodology was HLATools User Manual 161 of 172 HLATools_IVDManual_v1 1_Rev 0 pdf NIH Score NMDP code Normalization sound to correct for background noise or contamination and to normalize raw data readings The actual nature of a negative control may vary from product to product In LABType a probe that will not bind to any HLA sequence e InLABScreen a negative control serum that contains no HLA antibody accordi
184. state that the application is Unable to retrieve the revision date All of the cells in the table are editable fields to allow user updating and annotation Figure 7 10 Navigating to an Allele in the Serological Equivalents Table Allele to Serological Equivalent SerologicalE quivalent BestGuess B 3924 CET O EA sE BBlank Name B 3529 T_ h B al EU CN EEN ssa E o B The columns in the serological equivalents table contain the following information e Name the allele type and subtype assigned by OLI HLA analysis e SerologicalEquivalent many alleles have identified serological equivalents These equivalents are displayed in this table Null alleles i e alleles whose molecular variations are not expressed as changes in surface antigens are indicated in the Serological Equivalent Table as Blank where represents a serological type Allelic types that have been identified on the molecular level but not serologically are indicated as e BestGuess this column is currently unused Bw4 Bw6 or Cw1 Cw2 the Bw4 Bw6 and Cw1 Cw2 systems are biallelic antigen systems closely associated with HLA B and HLA C The Allele to Serological Equivalent controls are identical in function to those in the Product Information view with one addition Export XLS exports table in Excel compatible Comma Separated Value format Because the file includes Excel compatible metadata it cannot reimported into
185. stomization HLATools LABType Interactive interfaces contain a number of user customizable features that are not immediately apparent to the beginning user Some of the more useful features are described in this Appendix Modifying Tables The appearance and functionality of many tables in LTI are user editable By rearranging the column headers you can make the table present the data in different ways that may be more suitable for the way you work Changing Sort Order or Grouping The Closest Reactions subview is an example of a table that can be reconfigured to change the way the entries are sorted and grouped By default the entries are sorted by bead number Figure 5 13 To reconfigure the table so that it sorts the False Reactions by Allelel candidates drag the False Reactions header from its position at the top of the table and drop it beside the Allele2 header Then drag the Allele1 header and drop it at the top The reconfigured window will resemble that shown in Figure A 1 The reconfiguration is local applying only to the current table and is not persistent Version 1 1 105 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Figure A 1 Closest Reactions Sorted by Allele1 Match E Allele1 DPB1 04UHY 1 item Allele2 FalseReactions DPB1 1401 3 FN z Allele1 DPB1 1401 2 items Allele2 FalseReactions DPB1 5901 3 FN DPB1 6801 42 FP Allele1 DPB1 3501 1 item Allele2 FalseReactions
186. sword combination in the Create New SQL User dialog Figure C 12 Version 1 1 147 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure C 13 Creating a New SQL User HLA Tools Create New SOL User eS x New User Information User Name Marge Password p T Blank Password Goal Cancel 2 Launch the Connections tool and create a new connection to the other server using the SQL username password combination In this case we will call the new connection Bart and link to the server named starfish Figure C 14 Creating a New Connection for SQL Authentication Database Settings xi Connection Name Bart 1 Select or enter a server name starfish ONELAMBDA y foe 2 Enter information to log on to the server Use Windows NT Integrated Security Use a specific user name and password User name Marge Password z Blank Password 3 Select the database on the server LTI e Attach a database file as a database name pee les dt rn _ Using the Filename ee E Set Cancel 3 Enter the same username password combination used when creating the SQL user on the starfish server Figure C 14 4 Click Test Connection to verify the connection and Set to commit the settings and exit the dialog This SQL user will maintain access to the LTI database on the specified server until the connection is deleted from the Connec tion Names list in the Connections tool
187. t A sample of this information is shown in Figure 7 4 To update information for a LABType product 1 Access the listing of new One Lambda products at http download onelambda com pub tray_info Windows HLATools Labtype_Interactive 2 The product filename must have a LabTypeProduct extension as shown below Products designated by an R revision number have been retired If you do download a retired product it will appear in the HLATools Product Information Table but it will not appear in the Lambda Explorer Product Select pull down listing Figure 4 4 since its Visible attribute is automatically set to 0 See discussion on page 74 Tuesday Tuesday Tuesday Tuesday Tuesday Tuesday April April April April April April Figure 7 5 Selecting a One Lambda LabType Product to Update To Parent Directory 96188 94398 97245 97780 101928 104193 R550144 002 R7 LabTypeProduct RS5014 003 RS LabTypeProduct RS5014 004 R3 LabTypeProduct RS5014 005 03 LabTypeProduct RS5014 006 03 LabTypeProduct RS5014 007 02 LabTypeProduct 3 Click on the filename hyperlink to launch the updating utility A conventional Windows File Download dialog will appear To download the new product information directly into the LTI database select the Open option An Open With dialog will prompt you to select an application to carry out the download operation Select GenProd Version 1 1 OLI UD 0157 062006 75 of 172 HLATools_IVDMan
188. t Field contains application generated sample analysis messages which are also included in many of the sample reports this is a user editable field user added comments will be included in the reports Comments of the following sort are displayed here e Warnings alerts of low bead count and low positive control values when applicable e No False Reactions count of the number of MRPs in the sample and the allele assignments for each MRP expansions of ambiguity codes are also provided e Close Reactions count and listing by bead of possible False Positive and False Negative bead reactions that would yield additional MRPs e Modified Beads listing of beads whose cutoffs have been modified by the user 14 Previous Next navigation buttons to move to adjacent samples It is possible to jump to any sample in the batch by selecting it in the batch sample file tree 15 Accept commits the current sample assignments and comments including any user edits to the database and locks the sample from further edits Clearing the Locked check box re enables the Accept function When using the option that requires that samples be accepted and reviewed by two different analysts rather than one the Accept button changes to read Review after the first analyst has completed his or her review The sample is Locked after the first acceptance has been completed and cannot be unlocked until the second review has been completed Changes
189. t folders is Do not share this folder Select the Share this folder option Figure C 16 Luminex Output Files Workflow CAMyBatches Output Luminex 100 Analyzer Network Server AAA HLA Tools User 4 The default user group specification in the Share Permissions interface is generally Everyone Permissions levels are e Full Control allows the user to change file permissions take ownership of files and perform the actions granted by the Change and Read permissions e Change allows the user to overwrite files change file attributes and view file ownership and permissions e Read allows the user to read files in the folder view file attributes ownership and permissions Version 1 1 151 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Figure C 17 Output Folder Properties Sharing Output Properties 5 Select the Everyone group icon and clear the Full Control and Change checkboxes making sure the Read check box is checked Now everyone will be able to read or copy Luminex output files from this folder but not change or delete them 152 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure C 18 Share Permissions for C LuminexFiles Permissions for Output i 2 xi Share Permissions Group or user names Add Remove Permissions for Everyone Allow Deny Full Control Change oO o Read Cancel Apply To simplify
190. ter 3 Chapter 4 Chapter 5 Chapter 6 Uninstalling HLATools 1 0 2 0 0 teens 22 Complete Uninstall of HLAToo S o0ooooooorncrrnorronanm 22 Updating HLATools 14 ci0ctsacendieoedds de dr dd 23 lop AA FS 26 LABType Home Product Inserts 02 a IS A sd 27 Patient Worksheets soscscirarter cata dra 21 Bead Probe Information IA 27 Resolution Limitations 2 dr ias rie lie 28 Lambda Explorer Accessing Luminex csv Files 2 2 cics 0ssabncsbavekbbetbavsdbasebavauas 29 Importing Luminex csv Piles varias dis e e as E 31 HLATools LABType Interactive Synopsis of LTI Batch Analysis Views 0 0 0 e ee ee ee eee een eee 35 Batch Data Analysis cerce 252 2904 sSGendeeebde deter pert en e 36 Batch Results View io 22 lt 80udsbeuuwes bee kasd aiiai a Sa a ies 37 Bead Analysis VieWerscrnrogaci nes deedaheeverciesatdidciers dais 42 Control Values View Positive Controls 0 0 0 0 e ee eee eee 44 Control Values View Minimum Bead Count 04 45 Synopsis of Sample Analysis Views and Subviews 02 eee eee 46 sample Results Analysis conaordarasi dE ENON SEY weed oud abn 47 Assignments View with Matched Reaction Pairs and Close Reactions 47 Matched Reaction Pairs Subview 0 0 eee eee eee 52 All Alleles SUBVIEW asicirodtacinriora raros ida a ri da da 53 Type Subtype SIDE es de cand ede eahee Sere aer 54 Closest Reactions SUbViewW 2225 citar 55 Specificity SUBVIE
191. teractive Configuration Suite lol xi Po Connection Name gt File p Analysis Comments II Configuration E Database AttachiDetach ba Backup Restore pu Clean Update Local 2 Diagnostics User Management Modify Add OK Cancel Apply 7 Click Add to create a new connection Figure C 6 8 Inthe Database Settings dialog that appears Figure C 7 enter a name for the new connection This name can be an alias for the target computer to which you are establishing the connection or any name you wish Here we will create a connection named Homer 9 Click Set to OK the newly added name The new name will now appear below the Local connection in the Con nection Name list Figure C 6 142 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure C 7 Database Settings Database Settings xj Connection Name Homer 1 Select or enter a server name swordfish ONELAMBDA y 2 Enter information to log on to the server Use Windows NT Integrated Security Use a specific user name and password User name Password F Blank Password 3 Select the database on the server LTI Attach a database file as a database name Using the Filename Test Connection Set Cancel 10 The name of the server to which LTI currently has access will appear in the topmost editable field 1 Note that the name of the newly created connection Homer app
192. tes that the patient is heterozygous at the locus both columns will contain a different allele assignment if homozygous only the Allele 1 column will contain an entry if no assignments can be made both columns remain empty The definition of the NMDP or local code for each Allele is viewable in a tool tip Comment displays the first line of the sample specific comments and diagnostics specifically any warning or alert generated by the program such as Low Bead Count or Low Positive Control or notification of an ambiguous assignment see comment to the Tool Tip above 10 Requester the name of the institution requesting that the sample be tested Patient ID part of the patient information that can be entered and edited in the Patient Information Sub view see notes to Sample ID above Lek checkbox displays whether the sample allele assignments have been accepted further changes in allele assignment or edits to patient information or sample and patient comments can be made only if the sample is unlocked in the Sample Assignment subview Figure 5 8 Allele assignments that have been Locked will ap pear in the analysis reports as Corrected Relationship Family patient information that can be entered and edited in the Patient Information Sub view A 1 A 2 type assignments for Allele 1 and Allele 2 Sero 1 Sero 2 serological antigen equivalents for Allele 1 and Allele 2 Bw 1 Bw 2 Bw4 Bw6
193. ting blue or white color except for those with ambiguous assignments e Red sample needs review contains possible false reactions or is empty MRPs are possible by flipping bead reaction states as explained in the Tool Tip discussion above 10 38 of 172 OLI UD 0157 062006 6 6 06 HLATools_IVDManual_v1 1_Rev 0 pdf Orange ambiguous assignment which means that more than one allele assignment can be made to one or both alleles based on the observed sample reaction pattern The current bead combination does not provide suf ficient resolution to discriminate between two or more possible allele assignments Typical ambiguous states are when an MRP would require a false reaction or the suggested assignments for an allele do not agree at the subtype level Columns Sample ID until a unique Patient ID is assigned to the patient the Sample ID also serves as the Patient ID multiple samples in one or more batches may be associated with a single patient with a unique Patient ID BatchID the number assigned to the batch in the LTI database this is not the session or batch identifier in cluded in the Luminex csv file Well well numbers and X Y grid well location on the tray SamplelD the number assigned to the sample in the LTI database this is not the same as the Sample ID Allele 1 Allele 2 allele assignments for the paternal and maternal alleles at the locus in no particular or der if the sample indica
194. tool to add and remove users from the second server s list of users authorized to access its LTI database Assuming we are using Quimby s computer of the previous example skipjack which we will now call the Remote server we can edit the user s list on the target server swordfish as follows 1 Launch the User Management tool By default the users authorized to access the local LTI database are listed in the main panel The names of the local server and the SQL server instance are displayed at the top of the panel Figure C 11 Listing Local Server Users HLA Tools Interactive Configuration Suite a iol xj File x Analysis gt skipjackXONELAMBDA Comments II a List Users BUILTIN Administrators Configuration HIL Connections ne Database network1 Lisa Attach Detach network 1 Smithers Backup Restore network14Quimby Clean Update Merge Diagnostics User Management Add SQL User Add Windows User Group Delete User 2 To change the connection to the target server select File gt Connection gt Quick Connections gt Homer 146 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure C 12 Connecting to a Remote Server HLA Tools Interactive Configuration Suite ad Local Ctri L Custom Connection List Users Connectivas skipjack Guest b network SLanny E Database network Lisa Attach Detach network Smithers Backup Restore network Quimby Clean Update
195. types 77 c Cell death detection 157 Character encoding type specifying 109 Class I II proteins 156 Clean Database need to run 27 using all cmd script 136 Close reactions illustrated 158 Code point 109 Codes amino acids 155 Local XXn format 46 120 NMDP 120 162 nucleotide 163 nucleotide triplets 160 Color coding in Raw Data table 63 in Reaction Grid 57 in Sample Data bead profiles 40 Comments Editor HLATools Comments Editor tool 47 107 Complement 158 Corrected typing HLATools User Manual Index 167 of 172 HLATools_IVDManual_v1 1_Rev 0 pdf same as Accepted typing 51 Count Type 71 158 CREG cross reactive group 158 Cutoff Sensitivity 40 71 D Data sheets Bead Probe Information 73 Resolution Limitations 73 Data transfer file export type 108 Data Type means and median 71 Database Log Files 160 Primary Data Files 164 Database Login when printing reports 116 Date formats changing 107 E EB ethidium bromide 157 EDTA ethylene diamine tetraacetic acid 159 ELISA 165 Epitopes private and public 159 Excluded beads background color in Reaction Grid 40 57 63 Export type Comments editor 108 data transfer file 7 108 F False Negative defined 159 False Positive defined 159 False reactions 159 Figures see also Tables All Alleles Table 51 generating 124 Allele Symbols in the Reaction Pattern Histogram 50 Assignments View w Clos
196. ual_v1 1_Rev 0 pdf Figure 7 6 Downloading One Lambda Product Information File Download x You have chosen to download a file from this location RA55020B1_004_0c from download onelambda com What would you like to do with this file Open this file from its current location M Always ask before opening this type of file Cancel More Info 4 One of two log messages will appear e Ifthe selected revision of the product information is already in the LTI database a message will confirm the fact and the window may be visible for only a second or two The existing version will not be replaced e If the product is not present or needs to be updated the download should take about fifteen or twenty seconds 5 Alternatively you can Save the file to your computer You might choose the Save option if you want to update the product information at a later date or if you need to transfer the data file to a second computer that is not attached to the Internet When you Save a file please note the extension associated with the saved file If the updating utility is correctly installed on the computer and the file extension is correct the File Download dialog should display the icon shown in the circle in Figure 7 6 You can load the updated product information into the database just by double clicking on the saved file To confirm that the new revision of the product information has been imported into the LTI database l
197. umn Order p 106 700f 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Chapter 7 Reference Files and Their Maintenance HLATools LABType Interactive draws much of its reference information from three principal sources One Lambda product data sheets molecular allele serological equivalence tables and the NMDP allele code listing The HLATools Configuration Suite which is installed along with the main HLATools software modules provides tools that make it easy to review and update these three sources of reference information Survey of LTI Data Resource Files To access the HLATools Configuration Suite dialog Figure 7 1 select Start gt Programs gt One Lambda gt HLATools Configuration Suite gt HLA Update From within this dialog you can perform the following functions e File gt Connection menu option establish connections to databases on other servers see Connecting to a Re mote LTI Database p 138 e gt the label at the top of the dialog displays the name of the computer on which the currently attached database resides the SQL server instance associated with the database and the name of database in parentheses Version 1 1 71 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Figure 7 1 Accessing HLATools Reference Data Files E HLATools Configuration Suite ma x gt 001883 SEB OLD ONELAMBDA LTI Products Comments
198. user and laboratory specific custom reports Chapter 9 HLATools Legacy Reporter provides an overview of the different types of preformatted sample patient batch summary and special reports available to the HLATools user Appendix A LTI Interface Customization describes some techniques that users can employ to change the appearance of charts and tables in LTT Appendix B HLATools FAQ addresses a number of Frequently Asked Questions Appendix C Network Installation of HLATools deals with establishing user access in different configurations such as standalone workstations peer to peer installations and multi user network environments These instructions deal with some common scenarios that you may encounter when using LTI in a multi user laboratory environment A small Glossary of terms pertinent to HLA typing and screening is included Version 1 1 11 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf The printed manual concludes with an Index This manual has been designed to be printed out or viewed online The distribution media contains electronic versions of this manual in PDF and HTML format Viewing the HTML Version of this Manual When viewing the HTML version of this manual the Internet Explorer browser may repeatedly generate a message warning that the files contain active content You can suppress this message by reconfiguring Internet Explorer as follows 1 Select Start gt Settings gt
199. uthentication are available to control access to the local LTI database A SQL Users remote users are granted access by means of a username password combination different from that used for Windows network security See Linking to another Server using SQL Authentication p 147 B Windows Users this option employs standard Windows network security Anyone on your Windows net work can be granted access to a target computer s LTI database after he or she has been added to that com puter s authorized users list In this example we will establish permissions for a remote user named Quimby Click Add Windows User Group to access the like named dialog Figure C 4 Figure C 3 Simple Network Installation with two Users connection Quimby data from skipjack DB connection Homer data from swordfish DB server skipjack server swordfish database LTI database LTI local user Quimby local user Homer remote user Homer remote user Quimby Version 1 1 139 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Figure C 4 Users Groups on Windows Network HLA Tools Add Windows User Group xj User Group Name network Quimby Add Cancel Domain Local Existing Users List Users Select item to add Users C Groups 4 Make sure the Domain tab is active The Existing Users list should already be populated You can refresh the list at any time by pressing List Users to l
200. utoff sensitivity Very Positive Positive Between 100 and 110 of cutoff False Positive Overrides any other positive color assignment False Negative Overrides any other negative color assignment Between 90 and 100 of cutoff or bead has been used in False Reaction assignments involving multiple beads Negative Readings much lower than 90 of the cutoff value often provide insight into the level of background noise in the batch Very Negative oun 1 Current Bead ID number 2 lt gt use these navigation buttons to move to the adjacent bead alternatively jump directly to any bead in the assay by double clicking on the bead ID number in the bottom panel 3 Exclude Bead from Analysis Checking this box removes the bead from analysis The bead will no longer be used in making assignments such as Close Reactions although it will still be displayed in the Reaction Pattern His togram In the Reaction Grid table the columns of excluded beads are indicated by a medium gray background Note that removing a bead reduces the resolution of the analysis The exclusion only takes effect if the Bead Anal ysis view is unlocked 17 4 QC Data readings were collected by analyzing 96 individuals taken from the general population and generating profiles for each bead By default the Sample Data is shown in the main display when the Bead Analysis View is accessed Clicking on the QC Data thumbnail
201. utoff values and accepting suggested allele assignments Version 1 1 7 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_ Rev 0 pdf Patient Explorer a utility that collates multiple sample results collected from among one or more batches with a single patient identified by a unique patient ID Lambda Reporter a report generator that creates a variety of preformatted batch sample and patient reports The Lambda Reporter also exports special reports such as ABDR reports as well as files in xls txt pdf and doc formats HLATools Configuration Suite The HLATools Configuration Suite interface provides access to a number of utility programs used to configure the HLATools modules To access these programs select Start gt Programs gt One Lambda gt HLATools Configuration Suite HLA Update e Products modifies how LTT handles data on a per product basis Specifies type of mean and type of bead count used in data handling Specifies minimum bead count and positive control value thresholds Establishes order in which products appear in pick list when user matches a product to session data Specifies cutoff sensitivity used to classify strength of bead readings For details see Products gt View p 72 Update LABType product information from files downloaded from the One Lambda website For details see Products gt Update p 75 NMDP maintains NMDP ambiguity code listing Displays code list For details see NMDP
202. v 30 of 172 OLI UD 0157 062006 6 6 06 HLATools_IVDManual_v1 1_Rev 0 pdf Importing Luminex csv Files To illustrate file import we will import one of the sample files provided with the distribution From within the Lambda Explorer expand the C LuminexFiles folder and select ALot005 csv Figure 4 3 Selecting a File for Import Ea CA 7 29 2004 12 09 28 PM O 8 Samples 2 H E LuminexFiles Gy 041204BL62 csv 110004016104 3 i 08 12 04 PP1356 DPB Y 09 09 04 ASSO1BJ_0 augogwWorkshopALocusTest 10 08 04 PP1369 B _ BL6LowFirstS ample cs T BLOCUS lot6 csv RSSO1A_005 5 6 LabType A Locus Lot 005 1 BLot006 csv CLot003 csw i LJ DAB1Lot008 csy 123_104B_L6_1D1473 1 ligh_DR8_pli_96 csv Import KO When you select a Luminex csv file some items of header data from the file are displayed in the main panel of the Lambda Explorer This allows you to review the particulars of the input file without actually importing it The infor mation displayed varies with the version of the Luminex software that created the data file More extensive information such as product lot is recorded in files made with newer versions Figure 4 3 shows data from a Luminex 2 2 output file 1 Date and time when the Luminex run was carried out 7 29 2004 2 Sample size number of samples in the batch 8 3 LX100 serial number Luminex Reader serial number LX10004016104 4 Session ID assigned at t
203. value to the TABLE variable in the ALL bat batch file The default name for LTI databases is LTT However in the example shown in Figure B 13 the LTI database is called HUGO instead of LTT You can update a database with a non default name by running the batch file updateDB bat which is located in the C Program Files One Lambda HLATools Clean Database UpdateDB folder Before running updateDB bat you must be sure that the database file name there is identical to the name in ALL bat Figure B 14 ALL bat is located in the Clean Database folder All bat creates a new clean database The steps which the system executes when updating a database are briefly as follows The system 1 Creates a folder with the name Backup and a timestamp 2 Detaches the database 3 Copies the databases mdf main data file and Idf log file into the newly created backup directory 4 Renames the original mdf and Idf files by prefixing the word temp and appending the timestamp to the original filename 5 Attaches the newly renamed files 130 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf 6 Runs the All bat file creating a new database which contains the latest LABType product information NMDP and serological data 7 Runs MERGE which merges the temp copy of the database with the newly created database described in the pre vious step For detailed names of the files and folders involved in the u
204. ve cells Ethidium bromide a red dye binds to the DNA of lysed or dead cells and excludes the acridine orange A very strong positive reaction is indicated when over 80 of the cells are killed Reactions are graded using the NIH scoring scheme Table Gl 2 NIH Reaction Scores NIH Score Cell Death Rate Reaction Strength 0 None 1 1 lt 20 2 20 lt 40 4 40 lt 60 6 60 lt 80 Version 1 1 HLATools User Manual 157 of 172 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf Clean Database Close Reaction Code definition NIH Score Cell Death Rate Reaction Strength 8 80 lt 100 x unreadable Some typing sera register weak or moderate reactions if they react with an antigen in the same cross reactive group CREG as the main specificity The 0 8 scoring convention is also used in some non serological testing applications For example in LABType Interactive a positive test bead reaction is indicated by an 8 a negative reaction by a 1 The Clean Database utility creates the original instance of the LTI database It must be run before LTI can be expected to work Until it is run there is no database for LTI to use A database consists of two parts the Schema and the Data Clean Database builds the SQL Database Schema then fills it with some initial values list of One Lambda products serology table values etc Once this has been done there is enough of a framewo
205. w from further data entry patient data in all other samples associated with the patient are automatically updated 7 Locked clearing the checkbox enables the edit function 8 Relationship Family these entries are also included in the Patient Summary reports Bead Analysis Sample The Bead Analysis view is identical on both the sample and batch level See Bead Analysis View p 42 Reaction Grid The reaction grid consists of a header panel and a tabulation of the reactions of the beads for the given sample Figure 5 16 Sample Reaction Pattern Grid Sample G 3 DRAB1 11ARAP gt DRB1 110401 DRB1 110402 DRB1 1143 DRB1 1144 O Pin C DR811404 on A Go 6 13 Q DRB1 1428 DRB1 14 x12 DRB1 1431 DRB1 1439 DRB1 1411 DRB1 1410 DRB1 1404 x xX xX KK KK OM x xX KK KK OK OM 12 DRB1 114RAP DRB1 1146 x EA O xX KX OK OK DRBI 11UC DRB1 114PwWP DRB1 11JEN 13 DRB1 1109 DRB1 1150 DRB1 1118 DRB1 1125 x lt XXxxgpax x lt x xX xX X lt X XES x xX KX KK KK OK iB XK OK OK OK OK x xX xX KX KK OM OM ORS x xX xX KX KK MK XES Batch Results Assignments Bead Analysis 58 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf The reaction grid table Figure 5 16 shows the reaction pattern for the beads in the selected sample Reaction Grid Symbology and Background Colors e The observed reaction pattern for the sample is shown on the Rxn row e The c
206. with additional pairs of primers can be per formed Control beads intended to provide a reference value for a complete reaction Positive control values are typically used to confirm that the data collection methodology was sound and to normalize raw data readings The actual nature of a positive control may vary from product to product e In LABType a probe that will bind to all sequences for the particular HLA locus be ing recognized in the hybridization reaction e In LABScreen none positive reactions are defined in terms of cutoff values that are determined in different ways for the PRA Mixed and Single Antigen products e In Micro SSP a primer pair that amplifies a conserved region of the human beta globin gene the amplification product of the control primer pair verifies the validity of the PCR amplification A normalized value for a bead reaction that meets a preestablished cut off value The primary data file is the starting point of the database and points to the other files in the database Every database has one primary data file The recommended file name extension for primary data files is mdf See also Log Files 1df A DNA sequence that is matched by the probe on a bead The recognition site will usually be the exact reverse complement of the oligonucleotide sequence of the probe Recog nition sites are described in terms of the amino acids which the nucleotides encode rather than in terms of the nucleotide
207. xpanded ABDR by Batch Report Range e Allele Group Frequency Report e Allele Query Report lt under development gt e Reaction Assignment Report NIH score for each sample e Swiss Lab Report csv file intended for export only The Lambda Reporter allows the user to create reports with customized header and footer information generate reports in which the patient results are ordered according to a variety of sorting criteria and group analysis results for one or multiple patients from different test batches For overviews of the contents of the stock template formats see the corresponding report description in Chapter 9 HLATools Legacy Reporter Laboratory Report Header Information Laboratory specific information used by the Lambda Reporter is entered directly from the Lambda Reporter or using the HLATools Configuration Suite Figure 8 1 Laboratory Header Information Your Laboratory 1776 Your Street Your Town CA 91000 A PY Your Name Tel 800 555 5555 Fax 818 555 5556 E mail yname yourlab com www yourlab com To replace the One Lambda company icon shown in Figure 8 1 with your own laboratory company or institutional icon e Select HLATools gt Lambda Reporter e Select the Logo tab in the Search panel Figure 8 2 e Select the desired aspect ratio for the new logo Best results are obtained if the logos are the same sizes as indi cated on the Logo panel 170px x 27px for the wide logo and 38px x
208. y a single bead so they are also listed here in the Closest Reactions tabulation If the False Reactions in Figure 5 9 had involved two beads or more they would not appear here FalseReactions Allele1 B S514FKG FalseReactions FalseReactions FalseReactions FalseReactions llele1 Figure 5 13 Closest Reactions Subview FalseReactions 17 FP 3 items Allele2 B 5507 26 FP 2 items 42 FP 4 items 44 FP 2 items 49 FP 2 items Allele2 B 51AUX Allele1 B 5401 B 514UX B 5402 FalseReactions 53 FP 2 items Allele2 B 512Y 65401 B 512Y B 5402 FalseReactions 89 FP 2 items Al FalseReactions 94 FP 2 items E BYSLAFKG y B 510101 B 510103 B 510105 B 5103 B S111N B 5118 B 5127N B 5128 B 5130 B 5132 B 5133 oq r a JJ ad a a oO wv 4 lt D o un e Ler lt D a a o un M un taa 5 un un D pa a a T lt lt Collapse False Reactions Close Reactions one False Reaction are grouped by bead and reaction type False Negative or False Positive The default sorting mode of the subtables is by ascending bead number Other sorting modes can be specified by the user See the Modifying Tables p 105 for details on table customization and modification Cells display the suggested allele assignments that would result if the reaction status of a False Reaction bead were reversed T
209. ymous nucleotide substitution one that does not change the encoded amino acid and thus does not affect the antigenic expression of the protein on the cell surface or an intron polymorphism a mutation that occurs outside the region that encodes the amino acids in the protein on the cell surface Table B 2 HLA Allele Nomenclature Locus 1 2 3 4 5 Resolution DRB1 11 Low DRB1 T 11 01 High DRB1 11 01 01 01 High DRB1 11 01 01 02 N High Where 1 Allele type equivalent of serological antigen 116 of 172 6 6 06 OLI UD 0157 062006 HLATools_IVDManual_v1 1_Rev 0 pdf we YS NS Intron polymorphism Expression level of the allele Allelic subtype amino acid difference Non coding polymorphism e g synonymous nucleotide e A indicates Aberrant expression where there is some doubt as to whether a protein is expressed e C indicates an allele product that is present in the Cytoplasm but not on the cell surface e L has been shown to have Low cell surface expression when compared to normal levels e N has been shown not to be expressed e Q indicates Questionable expression in that the mutation seen in the allele has previously been shown to effect normal expression levels but the expression remains unconfirmed e S indicates that the allele codes for a protein that is expressed as a soluble Secreted molecule but is not present on the cell surface As of December 2005 per the

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