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1. 2 Select the preset for the slide or slides that you are scanning If you are using a CodeLink product that is on the list e g CodeLink 10K 20K select the name for the product This preset is applied to all of the slides when you load the cassette 3 Ifthe cassette is already loaded when you select the preset click Reset to apply the preset to all of the slides in the table 4 To apply a different preset to any individual loaded slide or multi scan slot choose the preset from the drop down list in the Scan table Preset column When you select a preset the options and values on the Channels Scan Area Image and Advanced tabs and the rectangles in the Slide View box change to those stored in the preset GppliedPrecision Chapter 5 Refining Scan Settings 51 Exposure Resolution and Sensitivity This section provides detailed information about the Exposure Resolution and Sensitivity settings to help you refine these settings It assumes that you know how to choose the settings in the Main Scan window For general instructions on choosing the settings see the appropriate section in Chapter 4 Scanning a Slide The CCD Camera At the heart of the scanner s optics is the CCD Charge Coupled Device camera The camera converts light from the slide into a digital signal that the scanner software processes to create the image file The CCD chip on the camera collects filtered light from the sample The light is filtered by the emis
2. Chapter 1 Getting Help 3 Document Conventions In order to make the information in this manual as easy as possible for you to locate and use the following conventions are observed Words in boldface are defined in the paragraph in which they appear Italics indicate emphasis e Bullets indicate items in a list or single step procedures 1 Numbered items are sequential steps for completing a procedure Note indicates information about the previous paragraph or step in a procedure Important indicates important or critical information about the previous paragraph or step in a procedure Tip indicates helpful advice See Also in the lower left corner of some pages lists cross references for the topic that you are reading AN WARNING Indicates important information regarding personal safety AN CAUTION Indicates important information regarding potential damage to equipment or software User Interface Description Conventions Boldface indicates the names of buttons menus dialog box options and fields Initial Capitals indicate the names of windows dialog boxes and tabs SMALL CAPITALS indicate the name of a menu command ALL CAPITALS SAN SERIF indicate the name of a key on your keyboard such as ENTER or DELETE Uniform width font indicates text to enter on a command line or in the GUI Definitions arrayWoRx Applied Precision s product family of Biochip Readers This consists of the original arrayWoRx scanner
3. To modify a preset 1 Select the preset that you want to modify in the list of presets Click the arrow to see a list of presets Select a preset from the list 2 Choose the options on the Channels Scan Area Image and Advanced tabs that you want to change 3 Make sure that the settings are correct The next step will overwrite the settings stored in the preset 4 Click Save 04 720046 000 Rev D 0804 50 arrayWoRx Standard amp Auto User s Manual To delete a preset Note You cannot delete a preset if it is being used by any of the entries in the Scan Table The software will display a warning message and prevent the deletion 1 Select the preset that you want to delete 2 Click Delete Scan Settings Preset Sample Savel Create Delete Are you sure you want to delete scan settings Preset Sample Click Yes to confirm that you want to delete the set 3 Click Yes in the Question dialog If you are scanning more than one slide or multi scanning use the Scan table to define specific presets for each scan To apply presets to slides You can apply custom presets that you have created ArrayWoRx also provides presets for CodeLink products 1 Click the Preset arrow to view a list of presets scan Settings Click the arrow to see Preset Bampez HE list of presets Entire Slide Benchmark ae Select a preset from the sample Sensitiv ample 1 x list Area C
4. intensity scale 68 Advanced tab enabling options on 38 Altitude 11 Analysis taskbar 20 Application stops responding 111 Applied Precision LLC contacting 6 Archive Manager CD 104 Archiving data to CD 74 Area Tab 81 Array See microarray arrayWoRx Auto about 4 loading slides into cassette 27 arrayWoRx Standard about 4 loading a slide 26 B Background intensity 147 subtracting 96 Bar Chart 99 Barcodes enabling symbologies 77 location on slide 57 using for file names 34 39 83 Batch of slides settings for 48 80 Benchmark scan area 48 Binning 51 Bright spots correcting 137 Brightness adjusting display 68 Buffer Underrun 138 Bulb See Lamp assembly age counter 77 life 130 replacing 130 resetting the bulb age counter 133 Button labels hiding and showing 74 78 Buttons Image Window 89 Main Scan window 78 task 110 04 720046 000 Rev D 0804 C Cable connections 13 Calibration Flat field 63 when to perform 63 Camera temperature 18 77 Cancel a scan 41 Capture screen 117 Cassette bails 28 eject 27 inserting slides 27 29 load into scanner 30 pictures of 28 removing from scanner 44 Cassette Idle Mode defined 34 to exit from 34 Caution defined 3 labels 14 CCD camera 51 checking temperature 18 description 9 linear response of 52 CD creating list of files on CD 105 mount and unmount 113 operating system support 10 save files to
5. 1 From the Main Scan window Scan menu choose Scan Options 2 Select Enable Extended Scan Area and click Done The scanable region of the Slide View changes to show the extended scan area Important When the Enable Extended Scan Area option is selected do not use full height barcodes These barcodes lift one end of the slide and cause a noticeable change in focus across the slide Use barcodes that are centered on the slide and are at least 2 0 mm from either side of the slide ppliedPrecision Chapter 5 Refining Scan Settings 59 Notes 1 When Extended Scan Area is not selected all requested scans are dimension checked If part of a slide is in the Extended Scan Area region a message is displayed and the scan request is cancelled 2 The Enable Extended Scan Area option is sticky It remains selected until you unselect it even if the scanner and software are turned off and on Using Deconvolution Deconvolution is a method designed to remove the small amount of blurring that is present in images obtained by all optical instruments Applied Precision s proprietary deconvolution algorithms preserve the amount of light throughout the image Blurred data is reassigned to its original location Use deconvolution to eliminate bright spot blooming Select this option when array spot sizes are bright or very large To set the Deconvolve option 1 Inthe Scan Settings area on the main scan window click the Image tab
6. 104 Centering image 89 Channels active channel for preview scan 81 adjusting intensity values 68 defining settings for 36 displaying intensity of 89 intensity ratios 97 intensity scale of 89 min and max intensity 55 number of channels to scan 81 properties displaying 84 save as TIFF 92 select colors for 93 setting filters and exposure for 36 show or hide 88 single channel scans 37 Channels Tab 36 81 150 arrayWoRx Standard amp Auto User s Manual Circle region of interest as 97 Cleaning the System 130 Clear selected slides 33 Clear Preview 75 Click 4 Close application 111 Close Main Scan window 21 CodeLink 50 Colors display 93 of lines in Image window 93 selecting color or grayscale mode 93 Computer workstation configuration 10 Connecting networks 10 Connection stitch panel 39 Connections workstation and scanner 13 Constrain to circle 97 Contact Applied Precision 6 Conventions document 3 Converting image file to TIFF 102 Copy file 112 Counter space for system 11 Counter bulb age 133 Create preset 48 Current rating of electric supply 11 Customer Service Hotline 6 Customizing Gnome settings 124 90 Cy3 Cy5 choosing filter set for 36 D Data saving for Linux and Windows 12 Data button 78 Data folder defined 48 opening a file in 67 Data Graph about 98 opening 70 Data Inspector 95 opening 70 using 96 Data Name 32 86 Data Table o
7. 5 using 5 Histogram about 100 of intensity distribution 97 settings 96 using with other tools 70 Hot Surfaces caution 14 Humidity 11 Idle mode cassette 34 Image centering 89 display attributes 94 information 92 152 arrayWoRx Standard amp Auto User s Manual moving in Image window 89 moving with scroll bars 89 setting intensity scale 90 Image file defined 25 displaying 67 Image tab 37 Image Viewer launching after scan 48 Image window 44 65 87 Info pages 109 Insert slide 26 Installation 11 Intensity adjusting display 68 adjusting maximum 88 and Percent of Optimum 55 background 147 90 displaying data 96 distribution 97 100 examining 70 frequency plot 91 normalize between panel edges 83 profile plot 95 ratio 97 statistics 97 viewing min max values 91 Intensity of lamp measuring 133 Intensity saturation 55 Intensity scale distribution 90 Intensity scaling 84 Intensity scaling display 90 Internet Address Applied Precision 6 K Kill App 111 L Label dimensions 57 Lamp about 9 access door 132 automatically turns on 26 intensity 133 scanner light 23 turning on 26 when to replace 133 Lamp assembly illustration 131 replacing 130 safety precautions 130 Launch Image Viewer after scan 48 Level of detail setting 56 Light bulb See Lamp assembly Lights scanner 23 Line profile 95 Linux 107 DHCP 12 network connections 12 static IP
8. Chapter 5 Refining Scan Settings 53 Resolution The resolution is the size in microns of the area on the slide that is represented by an image pixel The Resolution setting determines the binning method and thus the maximum signal to noise ratio Use Table 5 as a guide to choose a resolution setting to maximize signal to noise ratio and minimize the file size Note The higher the resolution the smaller the pixel size the ower the resolution the larger the pixel size Table 5 The Effect of Different Resolutions on a 30 mm by 20 mm Scan Area for a Two Channel Scan Resolution um Binning Signal to File Size noise MB Ratio 3 25 Higher 1 by 1 Lower 240 6 50 2 by 2 120 9 76 3 by 3 80 13 00 4 by 4 60 26 02 8 by 8 30 52 03 Lower 16 by 16 Higher 15 A good rule of thumb for choosing the resolution is to use a pixel size that is at least one tenth of the diameter of a typical spot or feature of interest For example use 3 25 for spots 40 um or less 9 76 for 100 um spots etc An asterisk in the Resolution drop down box indicates that the pixel size 5 10 15 and 20 um is interpolated Pixel sizes with no asterisk are based on the size of one camera pixel and the magnification of the objective 20 um camera pixels 6X objective 20 um 6 3 25 um These pixel sizes are determined by the binning method no binning 3 25 um 2 by 2 binning 6 5 um 3 by 3 9 756 um etc Interpolated pixel sizes are acquired at th
9. Click OK to close Sawfish configurator 15 Choose File Exit to close the Sawfish Control Center ppliedPrecision Chapter 10 Introducing Linux and Gnome 125 File Permissions File ownership and file permissions are the means by which Linux protects files from unauthorized use This section is a brief introduction to these concepts and describes how to view a file s permissions For more information about file ownership and permissions see Permissions under If you are new to Linux UNIX in the online GNOME User s Guide A file s owner is the user whose account was logged on when the file was either created or copied to the system The file s permissions are settings that determine which users can read write to or execute the file To view a file s permissions 1 Right click the file in File Manager to display the File menu Cee Just local softWoRx data sample_images f File Edit Settings Layout Commands Help aA P fee se Back Up Forward Rescan Home Icons Brief Detailed Custom f Location usr local softWoRxdata sample_images Poe S E games T Name info lmagqeFile2 Open Glib ImageFile2 man MyScan st Open with rpms MyScan_ yiow GS sbin MyScan_w Edit E Sy softwoRx MyTIFF_6 bin Samplelma Copy calibration_files Scale2005 Move to Trash config Scalez2005 E Sy data Scalez005 amp wm sample_images Scalezoos Move zScanFiles gt Symlink ta ch Th ___ Ees 2 1
10. Click to display parent directory See Also Only image files generated by an arrayWoRx scanner can be viewed in the Image window To view a TIFF file choose SHOW DATA FOLDER in the File menu to display the File Manager Double click the TIFF file in File manager to open it in a TIFF viewer such as The GIMP a Linux image viewer Note Image files have a file extension of stc To open an Image file 1 From the File menu choose OPEN IMAGE FILE arrayWoRxe Scan File Scan Utilities Help Open Image File Ctr O Show Data Folder Ctrl D Archive Data To CD Preferences 2 Select the image file that you want to open from the list of files The current directory is displayed in the Filter box Click in the Directories box to move to the parent directory Filter Box Selection usi ocal sonWoFndata sample_imagesitestt sid 3 Click OK Tip Another way to open an image file is to click Data Folder and then double click the file name in the Gnome File Manager Note To display a list of files with the extension stc replace the asterisk in the Filter box with stc and click Filter Saving Files to CD 104 04 720046 000 Rev D 0804 68 arrayWoRx Standard amp Auto User s Manual Scaling Display Intensities After you open an image in the Image window you can view features in more detail by adjusting the display intensity of each channel with the Intensity
11. For more about inactive presets see the online Help Chromatic Calibration Performs chromatic calibration for new filters If you would like to add third party filters to your arrayWoRx scanner contact Applied Precision Customer Service at 1 800 862 5166 or servicehotline api com Update Scanner s Software Opens Scanner Software Package Status that displays information about the versions of the scanner software and allows you to update the scanner software when you have an update CD Note Scanner Software Package Status displays information about the installed scanner software Click Help SOFTWARE VERSIONS to display information about the installed arrayWoRx software GppliedPrecision Chapter 7 Main Scan Window Reference 77 Eject Slide arrayWoRx Standard only Ejects the slide from the arrayWoRx Standard scanner Eject Cassette arrayWoRx Auto only Opens the cassette door and ejects the cassette from the arrayWoRx Auto scanner Help Menu The Help menu commands provide access to the online Help system and to miscellaneous information for both the scanner and software arrayWoRx e Scan File Scan Utilities Help S BB Slide View Overview of Scanning scanner Information software Versions Overview of Scanning Opens the Main Help window See Using Online Help on page 5 Scanner Information Opens a dialog that displays information about the system software and hard
12. Passwd Tasks Help ZILA Quit Passwd 5 Type the new password in Changing password and click Accept Changing password Changing the password for user awuser New UNIX password Accept Cancel Help 6 Click Quit on the Config tab to close LinuxConf Note Users can change their passwords by choosing PROGRAMS SYSTEM CHANGE PASSWORD from the Gnome footprint menu 04 720046 000 Rev D 0804 122 arrayWoRx Standard amp Auto User s Manual Deleting a User Account Before you delete a user s account decide what you want to do with the data stored in the account s directories You can either compress and store the data delete it or leave the data in the user s account directories where it is accessible only to a user logged on as root To delete a user s account you must first log on to the system as root and open LinuxConf To delete a user account 1 Ifthe Config tab in LinuxConf is not already on top click the Config tab 2 Inthe directory tree go to User accounts under Users accounts Normal This will display the Users Accounts tab 3 Click the user name in the Accounts list on the Users accounts tab Config Tab Config Contra Status Gomes 0e Information nen eee You must specify at least the login name A Networki r Networking and the full name Users accounts Normal User accounts User accounts Group definitions Change root password F The account is enabled Special
13. Table images of the scanned slides can be opened in Image windows for viewing using the File OPEN IMAGE FILE option on the Main Scan window This option is described in Displaying an Image File in the Image Window in Chapter 6 Viewing Image Data You can look closely at the image to determine whether the scan settings need to be refined and do a preliminary examination of image data before doing a complete analysis of the data ppliedPrecision Chapter 4 Scanning Slides 45 Figure 15 _ window eix File View w Tools w E w E a Zoom 4 0000 To zoom in or out in the Image window e To zoom in right click on an area e To zoom out middle click on an area See Also Displaying an Image File in the Image Window 67 Choosing Scan Options 48 04 720046 000 Rev D 0804 46 arrayWoRx Standard amp Auto User s Manual ppliedPrecision CONTENTS Choosing Scan Options Working with Presets Exposure Resolution and Sensitivity Displaying Pixel Intensity in Panel View Using the Resolution Calculator Slide and Microarray Dimensions Using Deconvolution Focusing the Scanner Flat Field Calibration 5 REFINING SCAN SETTINGS This chapter describes in detail how to make adjustments to the scanner and scan software in order to refine the settings described in Chapter 4 Scanning a Slide 04 720046 000 Rev D 0804 48 arrayWoRx Standard amp Auto User s Manual Choosing Sc
14. boot and then repeat Step 3 5 Type exit and press ENTER to continue the Linux start up process Note This procedure requires knowledge of the system s root password When the system leaves the factory no passwords are assigned We highly recommend that administrative passwords be assigned to the system at installation Avoid using the computer for doing other things while the recording is taking place Also you should have the files that you are archiving on disks that are local to your workstation Accessing a disk drive over the network may not work Record at a speed lower than the maximum for your recorder ppliedPrecision Chapter 12 Troubleshooting 139 Other Problems Cont d When Windows The system is waiting for If your system is connected to a local Windows network contact your is started a DHCP server that may system administrator If your system is not connected to a local Windows network use the following instructions to fix this problem workstation a l message 1 Start Windows Then right click the Network icon and select indicating Properties Windows is 2 Select Internet Protocol TCP IP in the Components list Restoring l i l connection to 3 Click Properties and select Use the following IP Address 172 16 197 1 is 4 Enter 172 20 20 21in the IP Address field displayed for 5 Enter 255 255 255 0 in the Subnet Mask field approximately l l l 90 seconds 6 Click OK When prompted clic
15. click Show 3D Graph Click Show Histogram Arrange the windows on the screen oe Sea Pe Select a region of interest ROI in the Image window to display its intensity values in 3D graph Histogram and the table in Data Inspector The display in each window changes automatically when you change the ROI in the Image window See Also Data Inspector 96 ppliedPrecision Chapter 6 Viewing Image Data 71 Selecting a Region of Interest in Data Inspector Pan Center Zoom Wheel Zoom Arrow The region of interest or ROI is a square region of the image that you can resize and move to visually examine image details This section describes one technique that you can use to select the ROI Follow these instructions and experiment with the tools to familiarize yourself with the ROI Figure 25 The ROI Containing a Single Spot Image Window 1 Samplelmage1 stc File View De aAa Tools ROI Box To define an ROI that contains a single spot 1 Open the image file 2 Zoom out until you can see the entire array in the Image window i Point to the zoom wheel in the lower left corner of the Image window ii Drag the zoom wheel up to zoom out down to zoom in 3 Center the region of the array that contains the spot in the Image window i Click 2 The pointer turns into a circle shaped icon when it is in the Image window li Click the spot in the Image window that you want to center 4 Zoom in until
16. now discontinued the arrayWoRx Standard scanner and the arrayWoRx Auto scanner 04 720046 000 Rev D 0804 arrayWoRx Standard amp Auto User s Manual arrayWoRx Standard The e scanner that provides manual loading and unloading of single slides array WoRx Auto The e scanner that has a 25 slide cassette with automated slide handling Mouse Terms Pointer The mouse pointer is the icon that indicates the mouse position on the screen It is usually a small black arrow but changes shape to show what effect using it will have For example the pointer turns into a small circle shaped icon in the Image window to indicate that clicking a spot will center it Point to Use the mouse to move the pointer to a position on the screen For example point to the File menu means to move the mouse pointer to the File menu Mouse button Mouse button refers to the left mouse button unless specifically stated otherwise For actions that require the middle or right mouse button the mouse button is specified For example using the middle mouse button drag the graph to rotate it Click Press the left mouse button and quickly release it To click an object on the screen point to the object and then press the mouse button and quickly release it Click refers to the left mouse button unless indicated otherwise For example click a file in the File Manager to open it means to point to the file press the left mouse button and qui
17. the bulb can explode If this occurs the fumes can be toxic Evacuate the room immediately and remain out of the room for at least 30 minutes Call Applied Precision for a replacement bulb assembly Overall steps for replacing the lamp assembly A Turn off the scanner and allow the lamp assembly to cool B Replace the assembly C Reset the bulb age counter in the scanner software D Perform a flat field calibration ppliedPrecision Chapter 11 Maintaining the System 131 A WARNING The lamp assembly can get extremely hot Make sure that the lamp is turned off and has cooled for at least fifteen minutes before replacing the lamp assembly A To turn off the scanner 1 Make sure that there is not a slide in the scanner 2 Turn off the scanner 3 Ifyou just turned off the lamp wait fifteen minutes for it to cool Note The lamp cooling fan will turn on automatically and stay on for a few minutes after you turn off the lamp WARNING DO NOT TOUCH THE BULB Fingerprints on the bulb can cause it to overheat when it is turned on and possibly explode If this occurs evacuate the room immediately and remain out of the room for at least 30 minutes Call Applied Precision for a replacement bulb assembly Figure 47 Lamp Assembly Lamp ca ble p pt a connector Te i w TN Lamp assembly thumb screws Ta e Finger tighten only Do not use a screwdriver Plastic knob CO Bulb 04 720046 000 Rev D
18. 0804 132 arrayWoRx Standard amp Auto User s Manual B To replace the lamp assembly 1 Open the lamp access door on the side panel of the scanner The lamp access door is the door closest to the rear of the scanner Lamp access door on side panel of scanner 2 Gently pull up on the door as you open it It should be horizontal when it is open You can also remove the door 3 Loosen the two lamp assembly screws at the top right and bottom left of the lamp assembly plate Note Ifthe plate is hot wait for it to cool before unscrewing the screws Grasp the plastic knob and gently pull out the lamp assembly 5 Disconnect the lamp cable from the assembly and put the assembly aside Remove the new lamp assembly from its packaging Note Use only replacement bulbs supplied by Applied Precision 7 Being careful not to touch either the front or back of the bulb refer to Figure 48 and check that the clips securely attach the bulb to the plate 8 Connect the lamp cable to the new assembly The cable connectors are keyed so that they only connect in the correct position 9 Install the lamp assembly Be careful not to pinch the cables 10 Hold the lamp assembly plate flush against the scanner frame and finger tighten the two screws clockwise DO NOT USE A SCREWDRIVER 11 Close the lamp access door ppliedPrecision Chapter 11 Maintaining the System 133 Figure 48 Check the Bulb Assembly before Installing It
19. 12 LinuxConf 119 Load cassette into scanner 30 slide arrayWoRx Standard 26 slides into cassette 27 Slide list 31 33 undetected slide 33 Load Eject button 44 Log on 18 Log on as root 119 Main menu Gnome 110 Scan 19 Main Scan window 73 closing 21 opening 20 Man pages 109 Measure Distances 95 Measure Features 69 Memory 10 Menu Main Scan 19 Microarray scanning through slide 26 size 34 79 Modify preset 49 Mount drives 113 Move file 112 Moving using scroll bars 89 Multi Scan and create file names from barcodes 83 button 31 of a single slide 33 remove all 34 using 33 N Network connection 11 connection for Linux 12 connection for Windows 12 default settings 12 DHCP 12 ports 12 Noise and spot detection 147 sources 147 ppliedPrecision Normal Scan 75 O Objective focusing 60 Online help 5 Open image files 67 74 TIFF files 67 74 Operating systems Linux 10 Linux network configuration 12 switch between Windows and Linux 18 Windows 10 Windows network configuration 12 Optimum Percent of 55 Options menu 94 Origin XY 81 P Panel defined 79 tool 79 Panel connection stitch 39 Panel flattening stitch 39 Panel View 83 about 84 colored bar 84 Panel Gnome 19 Password changing user 121 Password first time user 18 Percent of Optimum defined 55 Permissions file 125 Pixel coordinates and intensity values 95 displaying
20. 595 nm wavelength channel GppliedPrecision Chapter 8 Image Window Reference 91 Display Min Max Exp These values are the display intensity values of the dimmest and brightest display pixels in the Image window and the value of the exponent parameter respectively The exponent parameter determines how intensity values are distributed in the display The default values are the minimum and maximum intensity values of pixels in the image file You can refine the display by dragging the display intensity handles in Scale Image Tip To change the exponent value drag the lower portion of the intensity scaling function line directly above the Min Display Intensity handle Image Min Max The minimum and maximum intensity values of pixels stored in the Image file These values are actual data values from the Image file and cannot be changed Show Histogram Displays a frequency plot of the distribution of pixel intensities in the image file The y axis shows the number of pixels of a given intensity Log Displays the intensity histogram using a log scale The default value is 1 Done Closes Scale Image Restore Default Scale Applies default values to display scaling The default values are the minimum and maximum intensity values of the image file and an exponent parameter of one Help Displays the online Help topic for Scale Image 04 720046 000 Rev D 0804 92 arrayWoRx Standard amp Auto User s Manual File Menu The F
21. Be sure to load the slide face down so that the image is scanned through the slide OR e Ifyou are using arrayWoRx Auto insert the slide into the cassette with the barcode label facing down Note the number of the slot into which you load the slide so that you can select the appropriate slide to load from the Scan Table Insert the cassette into the scanner and load the appropriate slide using the Load Slide option on the Scan Table Click the Channels tab on the main window Select a CY3 filter and set its exposure time to 10 seconds Then click the Image tab and set Resolution to 3 2520 Click a to view a single image Then locate a bright speck on the slide and zoom in on that spot Choose Utilities Focus Scanner to switch to Focus mode Adjust the focus knob until the speck is in focus Then turn the focus knob 1 4 turn counter clockwise Stop the focus mode See Focusing the Scanner on page 60 04 720046 000 Rev D 0804 64 arrayWoRx Standard amp Auto User s Manual See Also 10 11 On the Utilities menu click CALIBRATE SCANNER Click Calibrate Scanner In Filter Set to Calibrate choose a filter set Cy3 for example Then select the Flat Field Calibration tab and click Calibrate Now Click Continue to start calibration When calibration for the filter set you selected in Step 7 is complete verify that Calibrated appears in the Calibration State field and that today s date appears in Last Calib
22. Bodipy TMR Lissamine Rhodamine LOLO 1 Lysotracker Red Mitotracker Orange PO PO 3 PO PRO 3 Resorufin Rhod 2 Rhodamine Phalloidin Rhodamine Red ROX 5 ROX R Phycoerythrin Sulfarhodamine 101 SYTOX Orange TAMRA 5 TAMRA TM Rhodamine TRITC Alexa 633 Alexa 647 Alexa 660 Allophycocyanin Bodipy 630 650 X Bodipy 650 665 X Nile Blue TO PRO 3 TOTO 3 To Be Released Alexa 568 Alexa 594 Bodipy TR Bodipy TR X Cy 3 5 Naphthofluorescein SYTO 17 Texas Red YO PRO 3 YOYO 3 1 8 ANS 6 8 difluoro 7 hydroxy 4 methylcoumarin 7 amino 4 hydroxycoumarin 7 amino 4 methylcoumarin Aminocoumarin Anthroyl Stearate Calcein Blue Cascade Blue Coumarin Granular Blue Hoechst 33258 Lysosensor Blue Lysotracker Blue Marina Blue Pyrene Quinine Sulfate True Blue 5 and 6 carboxy 2 7 dichlorofluorescein Acridine Orange Alexa 488 BCECF Bodipy FL Calcein Cy2 CyQUANT GR DNA Star FAM 5 FAM FITC Fluorescein Fluorescein Chlorotiazinyl Fluor X Lysosensor Green Mitotracker Green Neurotrace 500 525 Oli Green Oregon Green Pico Green Rhodamine 110 Rhodamine 123 Rhodamine Green Rhodol Green Ribo Green SYTOX Green TO PRO 1 TOTO 1 ppliedPrecision CONTENTS Example Two Scanners One Spot Image Noise Noise amp Spot Detection Background Dynamic Range Conclusion APPENDIX B SIGNAL TO NOISE For microarray experiments the signal to background ratio is calculated as the m
23. Gnome file manager More information about using the File Manager is available in the online Help To open the Gnome File Manager e Double click the Home directory icon on the desktop 5 Home directory O Or e Choose PROGRAMS FILE MANAGER from the Gnome Main Menu gt mE sf Rescan Home Icons Brief Detailed Custom La tpi 4 My Scan ste 3 sbin e Py G Sy softWoRx D LON t 3 calibration_files config H data ry ry 2 amp wa sample_images D D EN z5canFiles help Scale2005000_w6 Scale200500_w59 Scale200500_w68 Glib 85 tif 5 tif 5 tif Glog 2 A G out_files s A released_help Ca Bo Se E MyTIFF_685_595 Samplelmagel ste Scale2005000_w5 _xx tif 95 tif 125 267 476 bytes in 1 file Show all files The location of files on your system depends on its configuration The following procedure describes how to open an image file stored in the default data directory To open an image file 1 Open the Gnome File Manager 2 Double click shared _data 3 Double click the file in the File Manager to open it e stc files are opened by arrayWoRx e tif files are opened by the GIMP A file s Mime type determines which application opens it when you double click the file in File Manager See Launching Applications From the Gnome File Manager in the online Gnome User s Guide for information about Mime types GppliedPrecision Chapter 10 Introducing Linux and Gnome 113 To copy
24. Rev D 0804 20 arrayWoRx Standard amp Auto User s Manual Use the following procedure to open the Main Scan window To open the Main Scan window fp l Double click the arrayWoRx icon on the desktop or Gnome panel or Click the Gnome Main Menu button on the Gnome panel and choose PROGRAMS ARRAY WORX START ARRAY WORX wW CII amp Applications F utilities Doed oan L 2 Development Programs r amp Games amp Favorites I Pa Graphics Applets intenet amp KDE menus Gj Multimedia BA Run Settings a Panel System Er Documentation ock screen d Editors Lag out g ios W File Manager ps w Q Help system 8 wasview Start array WoRx Quit array WORX YO oe Y Y OF oy Gnome Main Menu button The Analysis taskbar will open Scan button 2 If arrayWoRx does not open click the Gnome Main Menu button on the Gnome panel and choose PROGRAMS ARRAY WORX QUIT ARRAY WORx to properly close the program Then repeat Step 1 3 Wait for the Power light to stop flashing and shine steady green then click the Scan button in the Analysis taskbar The Main Scan window opens If the scanner has not finished its initialization this takes about 2 minutes from the time the scanner is turned on the Main Scan window opens partially and waits until communication can be established with the scanner This partial window is normal The full window will open when the
25. Scan table If more than one slide is present a dialog opens that allows you to choose which slide to multi scan Once that slide has been selected it appears on a second and if applicable third fourth etc line in the Scan Table The Scan Table can contain up to 50 entries While arrayWoRx is scanning the file name of the current scan is displayed above the Slide View window When arrayWoRx is not scanning the number of the loaded slide is displayed Notes 1 The Multi Scan dialog is displayed only for slides that are detected by the system i e slides that appear in the Scan Table 2 When you multi scan a slide with Use Barcode For File Names selected arrayWoRx creates a different file name for each scan by appending a scan number to the barcode 3 When more than one slide is selected the Preview area is cleared between scans When performing a multi scan on a single slide with no other slides selected the Preview area is not cleared between scans 04 720046 000 Rev D 0804 44 arrayWoRx Standard amp Auto User s Manual Remove Cassette from the arrayWoRx Auto Scanner To remove the cassette from the scanner e Press the Load Eject button in front of the cassette door Load Eject Button Or e Click the Eject button in the Main Scan window File Scan Utilities Help Eject Button View the File When the scanner has finished scanning all the slides marked for scanning in the Scan
26. Scan Options Opens Scan Options that allows you to choose options for either a Preview scan or a Normal scan See Choosing Scan Options on page 48 04 720046 000 Rev D 0804 76 arrayWoRx Standard amp Auto User s Manual Utilities Menu The commands on the Utilities menu allow you to control the scanner They are generally commands that you will use to prepare for a scan or to maintain the system Scan array WoRx e File Scan Utilities Help am E Turn Lamp On Off Data Slide View Calibrate Scanner Previ Focus Scanner Set Active Filter Wheel Manage Inactive Presets Chromatic Calibration Update Scanner s Software Eject Slide Turn Lamp On Off Turns the scanner s lamp on and off The Lamp icon indicates whether the lamp is on or off Focus Scanner Opens Focus Scanner that allows you to start continuous acquisition so that you can focus the objective See Focusing the Scanner on page 60 Calibrate Scanner Opens Calibrate Scanner that allows you to perform Flat field and intensity calibrations See Flat Field Calibration on page 63 Set Active Filter Wheel Opens the Set Active Filter Wheel dialog box that allows you to select the active filter wheel after you have changed filter wheels Manage Inactive Presets Opens the Manage Inactive Preset dialog box that allows you change a preset so that it applies to filters in the active filter wheel copy a preset or delete a preset
27. Selecting channels 88 colors for wavelengths 93 scan area 34 Selector wavelength 88 Sensitivity setting 37 54 Serial number 77 Settings saving scan settings 48 Shared data creating subdirectories in 103 printing from 92 purpose of 12 saving to 101 Show or hide 3D Graph contours 98 tool tips 74 toolbar 94 wavelengths 88 signal variation 51 145 Signal To Noise Spot SNR 147 Signal to noise ratio 51 54 maximizing 52 Single panel acquiring image of 75 Size microarray 34 ppliedPrecision slide and microarray 57 Slide descriptions 40 deselecting all slides 33 40 determine if slide is detected 85 dimensions 57 graphical view 79 identifier 86 insert in arrayWoRx Standard scanner 26 insert in cassette 27 load or unload a single slide 40 manually load undetected slide 33 multiple scans of a slide 33 multi scan option 33 reset data names 34 scanning 25 select to scan 40 selecting a few slides 40 settings for batch 48 transparent vs opaque 27 type 57 unloading 41 viewing status 31 Slide View 79 clear images in 75 Slot 32 SNR sensitivity option 54 Software Package Status 76 Software version 76 77 Space requirement of system 11 Spot SNR 147 Start a scan 41 Start arrayWoRx software 19 Start system 18 Static See Electrostatic Statistics 97 Statistics Record file format 96 Status bar 42 86 Status light 23 stc file extension 39 Stitch panel connect
28. They Mean State Power Status Lamp Green Steady Ready slide loaded lamp on Green Initializing busy scanning warming up Flashing Amber Steady Standby cassette loaded system bulb use over 1000 hrs ready to load slide arrayWoRx Auto only Amber powering down slide loading unloading failed to light or Flashing bulb use over 1000 hrs Off power off no cassette loaded lamp off arrayWoRx Auto only Note All three lights simultaneously flash amber if a system fault occurs If there is a system fault press the Power button to restart the system If the problem persists contact the Applied Precision Hotline ppliedPrecision CONTENTS Handling Slides With arrayWoRx Standard Handling Slides With arrayWoRx Auto Use Preview to Define the Scan Area Choose the Filter and Exposure Choose the Resolution and Sensitivity Define the Advanced Options Start and Stop the Scan Scanning a Slide Multiple Times View the File 4 SCANNING SLIDES This chapter describes the basic steps required to scan slides For more detailed information about how to use the features of the software to refine the scans refer to Chapter 5 Refining Scan Settings To scan slides you insert them into the scanner and define settings in the scan software The system then acquires the images and creates image files An image file contains a table of intensity values and coordinates that describe excited fluorescence on the slide The image file ca
29. a Flat field calibration after adding filters replacing the lamp and every 200 hours of lamp use Cleaning the Calibration slide To ensure that flat field calibration works properly make sure that the calibration slide is clean Use the following precautions when you are cleaning the calibration slide Avoid getting the calibration slide dirty Handle it with care and store it in a clean dry place Do not use canned air or unfiltered house pressurized air to clean the calibration slide as this may contaminate it with fluorescent propellants oil droplets particulates etc Use a filtered deionized air source or a simple bulb blower to blow dust off the slide For stubborn dirt gently moisten the glass surface of the slide with your breath and carefully wipe the slide with a clean soft lint free cloth See Appendix A Do not use ethanol chloroform organic solvents or lens tissue to clean the calibration slide they will degrade its surface Only use a soft lint free cloth for cleaning this slide To perform a flat field calibration l Make sure of the following e The flat field calibration slide is clean e The system has been on for at least 20 minutes e The lamp is on and the indicator has stopped flashing Start the scanning software and open the Main Scan window Load one of the purple calibration slides into the scanner as follows e Ifyou are using arrayWoRx Standard insert the slide into the scanner
30. a user interface to control the scanner a CD recorder to archive data an external network connection and access to a Microsoft Windows environment through VMWare The workstation configuration is shown in the following table Table 2 arrayWoRx Workstation Configuration Minimum Workstation Configuration 1 6 GHz Intel Pentium 4 processor or better 1 GB RDRAM 64 MB video card 17 flat panel monitor 40 GB hard drive arrayWoRx Auto has an additional 80 GB hard drive 1 44 MB 3 5 floppy drive CD RW drive CD archival is supported only when using arrayWoRx software PS2 keyboard 3 button mouse RedHat Linux 7 1 Windows 2000 Scanning software on the workstation provides a user interface to the scanner giving you control of the excitation frequency resolution exposure and scan area Analysis software if installed performs quantitative analysis on the ratio and intensity parameters associated with each spot such as mean ratio standard deviation etc VMWare allows you to run Windows from the Linux desktop After you start Windows you can connect to a Windows network connect to a printer and install third party Windows applications just as you would in a typical Windows environment Note VMWare does not support CD archiving 1 e burning a CD To archive data to a CD use the arrayWoRx software in Linux AN CAUTION Installing any third party Linux applications on the arrayWoRx workstation voids
31. accounts Policies Login name Jawuser File systems Miscellaneous services boot mode Full name FO group awuse o Supplementary groups o Home directory opt home awuser Command interpreter opt ninbash S A User ID opt 500 Quit Act Changes Help Accept Cancel Del Passwd Tasks Help Base info Params Privileges Quit Del 4 Click Del at the bottom of the User Information tab to open Deleting account Deleting account pgilman You are deleting an account The home directory and the mail inbox folder may be archived deleted or left in place A Archive the accounts data x Delete the account s data w Leave the accounts data in place Accept Cancel Help ppliedPrecision Chapter 10 Introducing Linux and Gnome 123 5 Choose the appropriate option Archive the account s data compresses and saves data in the user s home directory to the Oldaccounts directory in the system s default directory Delete the account s data deletes all of the data stored in the user s home directory Leave the account s data in place deletes the user s account but leave the user s home directory on the system Only a user logged on as root can access the data 6 Click Accept to delete the account 7 Click Quit to close LinuxConf Creating a New User Account Creating a new arrayWoRx user account involves two steps A Creating the account in LinuxConf B Changing the window manager
32. during a scan Figure 11 The Channels Tab Area Channels Image Advanced No Channels EIRA Filter Exposure 16 evs oao 2 3 EYS oso s a ae oh 3 y TEEL j PETAS Active Channel 1 To define the channel settings for a scan 1 Click the Channels tab in the Main Scan window 2 Select the number of channels between 1 and 4 for the scan 3 Choose a dye name in the Filter drop down list for each channel The standard choices are e CY3 e CY5 e Alexa 350 DAPI e Alexa 488 FITC 4 Enter an exposure for each channel Click the Exposure field and type a value You can use the left and right arrow keys on your keyboard to move the cursor in the box Note The recommended minimum value for Exposure is 0 10 which is determined by scanner hardware GppliedPrecision Chapter 4 Scanning Slides 37 Choose the Resolution and Sensitivity Typical Settings Sensitivity High SNR Resolution 9 7560 Patent Pending The Resolution and Sensitivity settings give you control of the pixel size and dynamic range of the image generated by the scan Sensitivity determines the dynamic range or detail of information available over the range of intensities of the image See Sensitivity on page 54 Resolution determines the size in microns of the area on the CCD chip that is represented by an image pixel See Resolution on page 53 To choose the resolution for the scan Click th
33. in Panel View to see it more clearly see Figure 20 9 Watch Panel View and turn the focus knob labeled FOCUS incrementally until the image is in focus Wait for the image to be acquired between turns of the knob The Focus knob is very sensitive and only a small amount of rotation is necessary for fine adjustment Turning the knob clockwise moves the objective toward the stage Turning it counterclockwise moves the objective away from the stage 10 In Focus Scanner click Stop to stop image acquisition 11 Click Done to close Focus Scanner 12 Close the focus access door Figure 20 shows what dust on a slide looks like as the objective is focused Notice that the Percent of Optimum increases as the image becomes more in focus You can also track this increase in brightness using the intensity values displayed in Focus Scanner 04 720046 000 Rev D 0804 62 arrayWoRx Standard amp Auto User s Manual Figure 20 Dust on a Slide Used to Focus the Objective 1 Out of focus dust on slide 2 Dust specks look smaller as the objective is focused 3 Features become clearer and brighter 4 Image in focus sharp features and highest Percent of Exposure value See Also Choose the Filter and Exposure 36 ppliedPrecision Chapter 5 Refining Scan Settings 63 Flat Field Calibration Flat field calibration is an automatic adjustment that ensures that an evenly illuminated slide generates an evenly illuminated image Perform
34. intensity 89 distribution of pixel intensities 91 intensity distribution 97 intensity graph 97 setting 37 size 53 PMT scanner simulating gain 39 Point Values tool 95 Point with mouse 4 Position changing image position 72 moving image 89 on Slide view 86 restoring image to Home position 84 Power cable 13 Power light 23 Power requirements 11 Power up sequence 18 04 720046 000 Rev D 0804 Index 153 Precautions for changing bulb 130 Preferences setting 74 Preset button 32 Presets applying 50 applying to all slides 34 defined 80 deleting 50 80 modifying 49 options 48 80 resetting 33 Preview clear 75 exposure 48 scan 34 Preview button 78 Print area of slide 57 files 12 images 92 Progress indicator 86 Q Quit Main Scan window 21 R Ratio data 95 Ratios intensity 97 Recommended Resolution 56 Region of interest as a circle 97 defined 71 defining 97 intensity profile 98 selecting 71 size 97 Removing cassette 44 slide 78 Rename preset 80 Requirements installation 11 Reset bulb age counter 133 Reset button 34 85 Resizing scan area 35 Resolution 53 calculating 55 defined 37 82 guidelines 53 minimum 53 refining 53 154 arrayWoRx Standard amp Auto User s Manual relation to pixel size 53 Resolution Calculator 55 Right click 4 ROI See Region of interest Root user 119 Rotating TIFF image 39 S Safety 14
35. mouse to zoom up or down Zoom Down Makes the image in the Image window smaller 04 720046 000 Rev D 0804 94 arrayWoRx Standard amp Auto User s Manual Options Menu The commands on the Options menu allow you to change the appearance of the Image window File View Options Tools Display Ctrl D w Border Tools Allows you to hide or show the border tools The border tools are the buttons and scroll bars along the left bottom and right edges of the Image window When the border tools are hidden the display area of the Image window is larger Tip Hiding border tools is useful when you capture screen shots of images Display Opens Display Attributes which allows you to view and modify settings that affect how the image is displayed in the Image window Click Help in Display Attributes for more information about the settings Note You can specify values in Display Attributes to change or move the Scale Bar GppliedPrecision Chapter 8 Image Window Reference 95 Tools Menu The four commands on the Tools menu each open a dialog The descriptions below briefly describe each command For more detailed information about the tools in each dialog click Help in the dialog File View Options Tools E fi D wl Point Values Line Profile Data Inspector Measure Distances Point Values Displays the coordinates and intensity value of a pixel as you move the pointer over
36. numbered from one to twenty five from the bottom of the cassette to the top Slides can be placed into any of the slots and once the cassette is loaded the scanner will automatically determine the slot position of each slide Figure 8 The Cassette Rear Bail oH Z AN gt o _ _ Front Bail OX 6 AT Z e 4 J Front View KT K Rear Bail Cassette Guide Slot one on each side of cassette Side View Bails on the front and back of the cassette hold the slides in place while the cassette is outside of the scanner The front bail is moved to one side when loading slides into the cassette After the cassette has been loaded into the scanner the scanner opens the rear bails in order to load and unload the individual slides See Also Slide and Microarray Dimensions 57 ppliedPrecision Chapter 4 Scanning Slides 29 To insert slides into the cassette 1 Grasp the cassette in one hand and use your thumb or index finger to move the front bail to the side 2 Hold a slide by the end with the label or bar code and gently insert it completely into the cassette slot Ensure that the slide is placed into the slot so that it is parallel to the bottom of the cassette The scanner will not load slides that are cross slotted positioned so that one edge of the slide is higher in the cassette than the other Insert transparent slides into the cassette with the arrays facin
37. panel TIFF etc will use the bar code data in their filenames Enable Stitch Panel Flattening Uses multi panel data to refine the flat field calibration The image must contain a minimum of 16 panels in order to perform panel flattening Enable Stitch Panel Connection Normalizes intensities between the edges of panels Gain Allows you to simulate the effect of turning up the gain on a laser PMT scanner Like a laser PMT based scanner gain can introduce non linearities in low intensity signals and is sensitive to slide chemistry If you use a Gain setting to acquire images the images must be acquired with the same Gain and Exposure setting in order for the data to be comparable On the other hand data collected with Gain turned off can be compared even if different Exposure settings were used to collect the data We strongly recommend turning Gain Off for normal data acquisition Generate Images in TIFF Format Generates TIFF files automatically after the scan Image files stc are always generated after a scan and can be converted to TIFF at any time Choose this option if you want the image file to be converted to TIFF immediately after the scan Note that a separate TIFF file is generated for each channel For example one TIFF file is generated for Cy3 and another for CyS TIFF file data can also be flipped You can use combinations of the following three settings to achieve any end orientation e Flip Y mirrors the positi
38. scanner finishes initializing ppliedPrecision Chapter 3 Starting and Shutting Down the System 21 Closing the Main Scan Window Before closing the Main Scan window close any open scan dialog windows Notes 1 Do not close the Main Scan window by clicking the Close box in the top right corner of the window To ensure that the application closes properly use the following procedure instead of the Close box to close the Main Scan window 2 Closing the Main Scan window does not close any open Image windows or Image window analysis tools Closing the Analysis toolbar does close any open Image windows however To close the Main Scan window 1 Remove slides or cassettes from the scanner as follows If you are using arrayWoRx Auto and there is a cassette in the scanner click the Eject button in the Main Scan window or press the Eject button on the scanner and remove the cassette after the door opens Or If you are using arrayWoRx Standard and there is a slide in the scanner click the Eject button in the Main Scan window or press the Eject button on the scanner and remove the slide after it is ejected from the slot File Scan Utilities Help EEEIEE Eject Open Image File Show Data Folder Ctrl D Click Quit 3 Ifthe Save Scan Settings question box appears click Yes to save outstanding changes to the Preset that appears in the Scan Settings panel of the Main Scan window Click No to exit witho
39. settings for the new user account Important Note Do not create a new user account unless you know how to change the Sawfish window manager settings as described in this section If after reading these instructions you aren t sure how to change the window manager settings please call the Applied Precision Hotline for help A To create a new user account 1 Ifthe Config tab in LinuxConf is not already on top click the Config tab 2 Inthe directory tree go to User accounts under Users accounts Normal This will display the Users accounts tab awe_tech Linuxconf 1 19 subrev 2 Lis Config Control Status Users accounts You can edit add or delete users Networkin J Select Add to add a new definition E Users accounts Group definitions Change root password Special accounts Policies File systems Miscellaneous services boot mode games gopher Ip mail Quit Act Changes Help Quit Add Help 3 Click Add at the bottom of the Users Accounts tab to display the User Account Creation tab and the Base Info tab 04 720046 000 Rev D 0804 124 arrayWoRx Standard amp Auto User s Manual 4 On the Base Info tab complete the text boxes as appropriate The only required field is Login name Linux will assign the following values to fields that you don t complete Full name blank group username where username is the name that you type in the Login name field Home directory hom
40. the channel that is displayed and the minimum maximum and mean intensity values for that channel It also shows the intensity and percent of optimum values of the pixel that is under the mouse pointer The percent of optimum value is the intensity of a spot relative to the maximum intensity that the CCD camera can measure This value indicates whether pixels in the image are saturated Acquire Single Panel Image Acquires the image of a single panel at the location on the slide indicated by the Panel tool in the Slide View This provides the same results as clicking on the image in the Panel view or choosing Scan Get Single Panel from the main Scan window menu The image is acquired in the Active Channel the channel that is selected in the Channels tab Display Brightness Contrast Control Opens Panel View Scale Options that gives you control over the intensity scaling of the display in Panel View You can choose to have the scan software automatically scale the image or you can manually adjust the minimum intensity cutoff maximum intensity cutoff and the Gamma value These adjustments affect only the display in Panel View they affect neither the display in Slide View nor the data stored in the Image file Zoom In Make the image in Panel View larger You can also zoom in and center on a point by right clicking on that point Zoom Out Make the image in Panel View smaller You can also zoom out by middle clicking on a point Ho
41. this manual and the online Help system This chapter describes the conventions used in this manual and how to open the online Help system This manual provides information about the components installation operation and maintenance of the arrayWoRx Standard and arrayWoRx Auto Biochip Readers The online Help system provides the information included in this manual in addition to context specific information that describes the scan software s user interface 04 720046 000 Rev D 0804 arrayWoRx Standard amp Auto User s Manual Using This Book This manual is organized to help you quickly get started using arrayWoRx and to find more detailed information about the system when you need it Table 1 describes the organization of chapters in this manual Table 1 Organization of Chapters Chapter Describes 1 How the manual is organized and how to open online Help 2 The components of the system installing the system and safety 3 How to start and shutdown the system and open and close the scan software 4 The basic steps to scan a slide 5 More details about refining the scan settings 6 How to look at the image data generated from a scanned slide 7 The Main Scan window feature by feature 8 The Image window feature by feature 9 How to convert image files to TIFF and save data files to CD 10 An introduction to Linux and Gnome 11 Basic maintenance tasks 12 Error messages troubleshooting steps ppliedPrecision
42. 2 Select the Deconvolve Option To determine whether the data in an image is deconvolved 1 On the Image window choose File Get Info to open the Information dialog box 04 720046 000 Rev D 0804 60 arrayWoRx Standard amp Auto User s Manual Click Header Labels If the image data is deconvolved the Deconvolve Image SRC statement is displayed in the Header Labels window Focusing the Scanner Focus the scanner when scanned images appear blurry in the Panel View window To focus the scanner l 2 3 cf Turn on the system and start the scanning software Load a slide into the scanner Click the Channels tab and choose the following scan settings Channel 1 Filter Cy5 or the appropriate channel Exposure 0 3 and Resolution 3 252 Drag the panel tool in Slide View to find the feature on the slide that you want to focus on and to display its image in Panel View On the Utilities menu click FOCUS SCANNER to open Focus Scanner Open the focus access door on the side panel of the scanner The focus access door is the door closest to the front of the scanner In Focus Scanner click Start to begin continuous scanning The system will display panel images in Panel View ppliedPrecision Chapter 5 Refining Scan Settings 61 8 Drag the Panel tool in Slide View until the feature that you want to focus on is positioned in one of the corners of the Panel View image You can zoom in on the feature
43. 23 840 bytes in 1 file Properties Show all files Delete 2 Choose PROPERTIES to open the Properties dialog ImageFile2_w595 tif Properties Statistics Options Permissions File Permissions Current mode 0664 Read Write Exec Special user TT 7 _ seue croup T Seran omer 7 stew File ownership Group Jawuser BP oK amp Cancel 3 Click the Permissions tab to view the permissions settings for the file The settings are shown for the user who is logged on and using the system 04 720046 000 Rev D 0804 126 arrayWoRx Standard amp Auto User s Manual Changing the Workstation Time Date Before changing the time and date you must log on to the system as root and open LinuxConf If you do not know the root password contact your system administrator and have them perform the following procedure for you Note The time zone should not be changed at the same time the current time is set Close and open LinuxConf for each operation To change the time zone 1 Ifthe Control tab in LinuxConf is not already on top click the Control tab 2 Inthe directory tree click on date amp time The date and time options are displayed in an area to the right of the directory tree Click the zone button to open a list of available time zones Navigate in the list to the time zone that represents your local area and select it Click Accept Click Quit no check on the Control t
44. 57 1000 Phone 44 1672 518350 Fax 425 557 1055 Fax 44 1672 518359 Technical Publications email address techpubs apli com ppliedPrecision CONTENTS Scanner Overview Workstation Overview Installation Requirements and Recommendations Workstation Network Configuration Cable Connections 2 SYSTEM OVERVIEW The arrayWoRx family of Biochip Readers consists of e The original arrayWoRx scanner now discontinued e The arrayWoRx Standard scanner e The arrayWoRx Auto scanner The arrayWoRx Standard scanner provides manual loading and unloading of single slides whereas the arrayWoRx Auto scanner has a 25 slide cassette with automated slide handling The arrayWoRx scanners integrate a high performance CCD camera fluorescence optics a precision slide handling system and sophisticated scanner software The systems can scan microscope slides simultaneously capture the intensities of up to 4 fluorescence wavelengths on each slide and create data files One multi wavelength image file is created per scan and separate TIFF files can be automatically generated for each wavelength The arrayWoRx scanners can assist you in a wide variety of applications including cDNA or oligonucleotide expression arrays proteomics combinatorial chemistry antibody screening SNPs single nucleotide polymorphisms and genomics 04 720046 000 Rev D 0804 arrayWoRx Standard amp Auto User s Manual Figure 1 The arrayWoR
45. 8 arrayWoRx Standard amp Auto User s Manual Turning On the Scanner and Workstation When you first turn on the scanner you can perform scanner tasks such as focusing and previewing Before you can perform a normal full resolution scan the camera must cool down to its operating temperature If arrayWoRx displays a message that indicates the cameral temperature is too high wait until the message closes before you try to perform a normal scan Note The system ships without a Linux or Windows password We highly recommend that administrative passwords be assigned to the system upon installation To start the system 1 Turn on the scanner by pressing the power button on the scanner s front panel The Power Status and Lamp lights will light according to Table 3 on page 24 e scanner Lights Scanner Lights Power Button Power Button The arrayWoRx Standard scanner The arrayWoRx Auto scanner Turn on the monitor 3 Turn on the workstation Type the username awuser and press ENTER If a Linux password is assigned to the system type your password into the appropriate field and press ENTER again Linux and Gnome will start up Before you try to perform a normal full scan wait until the Power and Lamp lights stop flashing and turn steady green If arrayWoRx displays a message that indicates the camera temperature is too high wait until the message closes before you try to perform a normal scan 5 Double click the arrayWo
46. 95 W 685 _j met ae me Output Directory usrocavsoftWoRx data sample_ima File Prefix HiResSampi TIFF Options Output Size 24 bit RGB IT Scale TIFF Images Using Display Min Max Values Below Wave 1 Color G Min Max pisos B55340 Wave 2 Color R Min Max s1s08 p55340 Done Do It GppliedPrecision Chapter 9 Saving Image Data 103 3 To save a portion of the image in the Image window to TIFF click Select Region and select the region in the Image window to save lEz ss l Le 4 Ifyou want to save the file to a directory other than the one that appears in the Output Directory field click Output Directory and browse to a directory or type in a directory If you want to save TIFF files to a different directory we recommend that you create and use subdirectories folders inside the shared_ data shared drive 5 The software names the file based on the file prefix Change the name in the File Prefix box if you want to use a different prefix 6 Select the TIFF format by choosing an Output Size option 7 Click Do It to complete the export 04 720046 000 Rev D 0804 104 arrayWoRx Standard amp Auto User s Manual Saving Files to CD File Window The first procedure in this section describes how to save files on a CD R using the CD Archive Manager Choose CD RECORDING or ON WINDOW on the Help menu for a detailed description of the Archive Manager features Notes 1 The Archi
47. Check that the bulb is securely clipped to the assembly C To reset the bulb age counter Start the system Open the Main Scan window On the Help menu choose SCANNER INFORMATION to open Scanner Information Click the Advanced tab Click Reset Bulb Age Click Yes to reset ce Bulb Age counter to zero Click Done ot ee ee D_To perform a flat field calibration e See Flat Field Calibration on page 63 Measuring Intensity The intensity of the light that reaches the slide is a critical determinant of the quality of the images that you produce It is important that the intensity not vary significantly for each batch of slides that you scan Take periodic intensity measurements to make sure that intensity levels are within the acceptable range If the intensity level falls below the level shown in Table 8 for a lamp assembly in use or the lamp has 1000 hours or more of use replace the lamp assembly 04 720046 000 Rev D 0804 134 arrayWoRx Standard amp Auto User s Manual Click Calibrate Scanner Table 8 Acceptable Minimum Intensity Levels New Lamp Lamp Assembly in Assembly Use Cy3 3 0 1 5 Cy5 9 0 4 5 To measure the intensity l 2 3 Turn on the system and start the scanning software Open the Main Scan window Load the Intensity Calibration slide with the label facing down Important If you are using an arrayWoRx Auto scanner note the number of the slot into which you load the slide so that you
48. OMIESH Service HONE sas earch E EE SN cadena 6 COMOT OIC E arni ee em non E AE AEA AE 6 2 System Overview 7 Sanner OVEN ICW Tarere a rem eee rne meen eens es 8 IW OBKSLALIOM OVE IVIC Wea Cheater sticestie sa Gi samnd E E E EE E ETE 10 Installation Requirements and Recommendations cccccccccccceeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeees 11 Workstation Network Configuration cccccccccccccccccececececcceeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeey 12 LuxCon Noor AON oc oats cs ceecstetargiec a 12 Windows Conn aurat ON saesae e a a rs paeeartee helen 12 Cable Connec ON Sauce a eee RE E E E E eae 13 EES A DEEE EEA TAIF A E AEA TI AAE EE A E A EAA A AE T 14 Ultraviolet Leto tan CAL ineei a e a E e 14 Class 1 LED Product Intra ti O10 yc5s2 2soce 3a 05cbss2tyeasounsoxaheraub ip sacdeeiseaevamdounsoxetoravbtivoarselaeteee 14 Caution anid Warning La DENS me i5 0scacksachscananavdcanetesseandunnscdsaciugenvesav ei aiea ea Eaa reaa BREE 15 3 Starting and Shutting Down the System 17 Turning On the Scanner and Workstation cccccccccccccccccccccceceeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeees 18 Switch Between Linux and Windows ccccceeesessessssesseeeseeeeeeeeesesseseseessssesesessessseeseeegaaas 18 Opening the Main Scan Window ccccccccccccccccceceeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeee ey 19 Closing the Main Scan Window 0 0 sssssss0 se0sesedosseosenovevssnaesesvensbveseoedodeno
49. Quick Scale slider See Scale Image on page 90 Notes 1 The adjustments you make to the display intensity values do not affect the data stored in the image file 2 When the mouse pointer is on the image the Status bar displays the intensity of each channel for the pixel that is under the mouse pointer Figure 22 The Quick Scale Scroll Bar and Wavelength Selector File View Options Intensity Quick Scale Wavelength Selector oom 0 1667 To select a wavelength e Click the wavelength in the wavelength selector When multiple wavelengths are being displayed the selected wavelength is indicated by a yellow box To adjust the intensity scaling e Drag the Quick Scale Intensity slider to adjust the display intensity of all displayed wavelengths simultaneously Or e Press and hold the CTRL key while dragging the Intensity Quick Scale slider to adjust the display intensity of only the selected wavelength Note Drag the Intensity Quick Scale slider down to increase the display intensity up to decrease the display intensity GppliedPrecision Chapter 6 Viewing Image Data 69 Measuring Feature Dimensions Measure Distances allows you to measure the distance between two points in the Image window This section describes how to open the tool and make a basic measurement Click Help in Measure Distances to view detailed information about the tool in online Help Figure 23 Measuring the Diameter of a Spot with Meas
50. Rx scanner icon on the desktop to start the software Switch Between Linux and Windows The arrayWoRx software runs on the Linux operating system When you start the workstation the Gnome desktop appears However you can use the VMWare application to run Windows from the Gnome desktop You may need to run Windows to connect to your Windows network and to use optional analysis software e g Tracker ppliedPrecision Chapter 3 Starting and Shutting Down the System 19 To start Windows e On the Gnome Desktop double click the Windows 2000 icon VMWare starts and opens Windows VMWare opens the Windows operating system in a window on the Linux desktop You can typically switch between Windows and Linux by changing the position of the mouse pointer You can switch to Windows by moving the mouse pointer over the window that contains Windows and switch to Linux by moving the mouse pointer outside of that window However in some cases e g when you are shutting down Windows or using the Windows Full Screen option you may need to use the CTRL ALT key to gain control of the mouse in Linux To switch from Windows to Linux e Press CTRL ALT to switch control of the mouse from Windows to Linux or Linux to Windows Opening the Main Scan Window The Main Scan window provides access to tools that control how the scanner scans a slide Open the Main Scan window after you turn on the scanner and the workstation 04 720046 000
51. SE from the File menu instead of using the window s close box 2 Click the Gnome Main Menu button Programs Favorites Applets gr KDE menus B Run oF Panel Lock screen 4 Inthe Logout dialog choose Halt to shut down the workstation Choose Logout to log off of Gnome and return to the Login screen 5 Click Yes to log out of Gnome or click No to return to the desktop without logging out GppliedPrecision Chapter 10 Introducing Linux and Gnome 109 Getting Help on Gnome If you cannot find the information you need about Gnome in this chapter refer to the Gnome online documentation To open the Gnome online documentation 1 Turn on the workstation and log on to Gnome 2 Click the Help button on the Gnome Panel at the bottom of the screen Help Button Tip You can also open Gnome Help by middle clicking anywhere on the desktop and choosing Help GNOME HELP from the menu Figure 41 The Gnome Online Help Contents Page Gnome Help Browser el x Eie Window View Settings Help a eS ae oe Back Forward Reload Index History BMarks Help Location ftoc GENOME HELP INDEX GNOME Users Guide Man Fages Info Pages GNOME Documents The Gnome User s Guide is a useful introduction for new users and a basic reference for more experienced users The Man pages are a collection of documents that serve as a technical reference to Linux commands They are useful to e
52. Slide Status in Chapter 4 Scanning a Slide for additional information regarding this option Multi Scan Allows a single slide to be scanned multiple times during the same scan job using different presets or the same presets Disable Deselects all of the Scan toggle buttons so that none of the slides in the table are selected for scanning Reset Applies the selected preset to all of the slides in the table This option also removes all multi scans resets all slide data names and reselects each loaded slide to be scanned once Slot Allows you to see which slots in the cassette contain detected slides Scan Toggle button that determines whether or not the slide will be scanned Click the button to toggle the setting Blue indicates that the slide will be scanned gray indicates that the slide will not be scanned 04 720046 000 Rev D 0804 86 arrayWoRx Standard amp Auto User s Manual Preset Displays the default preset defined for the slide and allows different presets to be selected for each slide or multi scan Data Name Displays the slide identifier If bar code data is being read from the slides this identifier will be read from the bar code If not this field will display a slide name that corresponds to the slot name Custom names can be manually entered All files generated from the scanned slides e g panel TIFF etc are named using the identifier displayed in this field Note Data name lengths up to 70 char
53. a isnt cates castes causa oces meant EA Utilities WI CIs ie os seat E dap iedenclisantc eons sesel aAA He M 1 a eemeeeneee crore ecm er a Rar etn n Renee BONS eset sachet bi scinde eecwae a a E casos SS VIEW enen said orth ode Sead cee So ades EE A aA Scan Settings PROS CE iss ctor i E ee wales ete Scan Scttines PADS 2ien tate eee A Panel Vi Wenna te ended nant ante eee temeeasnten is ne OF 1 E 0 ge tent E CPT Co nD EE SE N ChesStaius Bahnen isleccecsen ts aa aise ede as Image Window Reference The Image Window Buttons cccceecccceceeeeeeeeeeeeeeeeeeeeeeeeeeees Seale TMa et Re nt rn ec ce eT ne gD Pile Meni 3355 8 orcas i datsesaddancce a aes Wil ys MEM aeiiae e E a ieaddsraneaataee OPNS MENU 95 30 chs Sasol tied hw ee eal eaa ene atid ae eeie Tool Meisi a a dees HeD MeD a a Re Data ISDE COR ena ON ID Daa rapie ea eek eee ee ee E E HSO oran ei A A A A A PS Saving Image Data OVEN OW ase a A A Converting an Image File to TIFF iirrainn is Savne oo Corse 0 Gentes netten ne rece stn nen nr i heen aan nen eer nee ene 10 Introducing Linux and Gnome BOAS id oh eee EN EE AAE E E TE mE ppliedPrecision Logging On to and Off of Gnome ccccccccccccceeeeeeeeeeeees Getting Help on Gnome ccc ccccccceeeeeeeeeeeeeeeeeeeeeeeeeeeeeeees The Gnome Desktop viiscs cece sataceniatiescecieiassheiaasehaseieieia einai Managing Files with the Gnome File Manager Working with the Floppy Disk and CD Drives Using t
54. ab to close LinuxConf ae ae To change the time and date 1 Ifthe Control tab in LinuxConf is not already on top click the Control tab 2 Inthe directory tree click on date amp time The date and time options are displayed in an area to the right of the directory tree 3 Enter the current hour information in the hour text box Guidelines for using Linux Following these guidelines for using Linux will help you to avoid problems with arrayWoRx Screensaver If the screensaver is on turn it off Screensaver activity can interfere with the normal function of the system particularly during long periods of data collection To turn off the Screensaver e From the menu in the task bar choose Programs Settings Desktop ScreenSaver Then select No Screensaver in the Screen Saver list on the Screensaver dialog ppliedPrecision Chapter 10 Introducing Linux and Gnome 127 Incompatible filename characters Mac and Windows OS filename conventions allow spaces or other characters that aren t safe or sanctioned by Linux The following rules apply to Linux file and directory names e Filename length supports up to 255 characters e You cannot use as a file name this is always the name of the root directory e Linux is case sensitive be aware of this when you are using capital and lower case letters e Many characters are not allowed in file names because they have reserved meanings These include
55. acters are supported The Status Bar See Also The Status Bar displays different types of information depending on whether scans are being set up or are in progress e While you are changing the scan area in the Slide View or defining a pixel size in the Resolution dialog the approximate file size required for the scanned image appears in the right side of the Status Bar This includes the stc file and any TIFF files to be created Status File Size 100 82 Mb e While the Scan dialog is open after you push the Scan button the Status bar displays the estimated scan time Status Scan time estimate 1 35 21 e During the scanning process the Status bar is a progress indicator that displays information about the status of the scanner The left side of the Status bar displays the progress of a scan The right side displays the percentage of the scan that is completed and the time remaining for the scan Status _ 45 Complete Time remaining 0 30 e Atany other time the left side of the Status bar displays the XY position of the mouse pointer when it is on the Slide view The origin is the bottom left corner of the slide area Status MY 40 81 0 34 Load the Cassette into the Scanner 30 GppliedPrecision CONTENTS The Image Window Buttons Scale Image File Menu View Menu Options Menu Tools Menu Data Inspector 3D Data Graph Histogram IMAGE WINDOW REFERENCE Menu Bar Window Num
56. ally turns on The lamp icon in the Main Scan window turns yellow and the Lamp light on the scanner s front panel turns steady green when the lamp is on and ready for scanning ppliedPrecision Chapter 4 Scanning Slides 27 Handling Slides With arrayWoRx Auto Insert the Slides into the Cassette Insert transparent slides into the cassette with the arrays facing down Insert opaque slides with arrays face up We do not recommend mixing transparent and opaque slides in the same scan job If you do you will need to perform a manual focus change and all transparent slides will need to be inserted with the arrays face up This can reduce the signal to background on the transparent slides by a factor of two Notes 1 If the Status light is shining a steady green or amber there is already a cassette in the scanner Click the Eject button on the Main Scan window or press the Load Eject button located in front of the cassette door to open the door and eject the cassette 2 If the Status light is off and the cassette door is closed the scanner may have entered Cassette Idle Mode see page 34 Press the Load Eject button to open the cassette door Figure 6 The Eject Button in the Software File Scan Utilities Help Figure 7 The Load Eject Button on the Scanner x Load Eject button 04 720046 000 Rev D 0804 28 arrayWoRx Standard amp Auto User s Manual The cassette holds up to twenty five slides Slots are
57. an Options This section describes the options in the Scan Options dialog box From the Scan menu choose SCAN OPTIONS to open the dialog box Figure 16 Scan Options E E an O ptions Choose Data Folder shared_data Enable Extended Scan Area M Launch Image viewer After Scan 7 Exposure Time For Preview Scan seconds on Done Help Choose Data Folder Opens Select File where you can choose the folder in which to store the files generated by the scan Note Directory path lengths up to 177 characters are supported If you enter a directory path that exceeds that length the path name is truncated Enable Extended Scan Area Increase the length of the scanable slide area to 73 6 mm This option also changes the minimum X dimension for scanning to 2 0 mm This option is available only for systems that have extended scan area hardware To determine whether your system has this hardware check the serial number Upgraded systems have serial numbers E6X XXX ESA or A6XXXX ESA New systems have serial numbers E65X XX 2 color E64X XX or A64XXX 4 color Launch Image Viewer After Scan When this option is selected and a single entry is enabled in the Scan Table arrayWoRx automatically opens the Image Viewer and loads the new image data If more than one entry is enabled in the Scan Table the Image Viewer does not open automatically Exposure Time For Preview Scan seconds Choose the exposure time for preview sca
58. arrayWok i Biochip Reader built with precisionware Standard amp Auto User s Manual Revision D ppliedPrecision arrayWoRx Standard amp Auto User s Manual Revision D of the User s Manual for the arrayWoRx Biochip Readers Part number 04 720046 000 Rev D 2004 Applied Precision LLC All rights reserved No part of this manual may be reproduced transmitted stored in a retrieval system or translated into any language in any form by any means without the written permission of Applied Precision LLC Information in this document is subject to change without notice ArrayWoRx is a registered trademark of Applied Precision LLC Cy3 and Cy5 are registered trademarks of Amersham Biosciences All other registered names and trademarks referred to in this manual are the property of their respective companies Applied Precision LLC 1040 12 Ave NW Issaquah WA 98027 425 557 1000 FAX 425 557 1055 For questions or comments regarding this manual please email techpubs api com ppliedPrecision Contents lil Contents 1 Getting Help 1 Cno EDS DOO Kane irs ee ete ees cade ean eee acca A ee 2 TOC WIS EAC Oly CONS sche Sa e a nea mua aie a 3 User Interface Descriptions Conventions sekusi a a a aaa aa aia 3 BrO a A E A E E A E O AA EEA TOE AT EO AA EEATT EE eee eee 3 Mo e Le esa a E E E Miedicade waht tinue 4 Cone Onnie Pe Dea a r E eee 5 Contactino A pplied Precom LEC iat scedecese ht a a A aE 6 CAIST
59. ber Button Channel Intensity Quick Scale slider Zoom Wheel Zoom to One Status Bar This chapter provides a brief description of each feature of the Image window including Data Inspector If you are looking for instructions on the use of a particular feature check to see if it is described in Chapter 6 Viewing Image Data You can also find detailed descriptions of each feature in online Help which you can view by clicking Help in the window or dialog for the feature you are using Figure 36 The Image Window Image Window 1 Samplelmage1 stc File View Options Tools Help E Scroll Bar Arrow Scroll Bar Slider Scroll Bars IY an Scale Bar 04 720046 000 Rev D 0804 88 arrayWoRx Standard amp Auto User s Manual Menu Bar The File View Options Tools and Help menu commands are described beginning with File Menu on page 92 You can execute some menu commands using a shortcut key The shortcut key for such commands appears next to the command on the menu For example to save a file you can either choose SAVE on the File menu or press CTRL S Window Number A number is assigned to each open Image window If you have more than one Image window open you can refer to a window by its window number You can also use a window number to specify input by dragging it into the Input field instead of using the file name Buttons Navigate the Image window and adjust the inten
60. can select the appropriate slide to scan in the Scan Table Then insert the cassette into the scanner and load the appropriate slide using the Load Slide option on the Scan Table On the Utilities menu choose CALIBRATE SCANNER to open Calibrate Scanner Scan array WoRx e File Scan Utilities Help e H Turn Lamp On Off Previ Focus Scanner Calibrate Scanner set Active Filter Wheel Manage Inactive Presets Chromatic Calibration Update Scanner s Software Eject Slide Set the following scanning parameters Cy3 channel selected with exposure of 0 1 sec resolution 3 25 um sensitivity High SNR Drag the Panel Tool the yellow square in the Slide View to different locations while checking the Percentage of Optimum value reported at the bottom of the Panel View window Find an area that has a value of 6 or less using the Cy3 channel and the settings described in Step 5 above Avoid areas with bright specks which have high Percentage of Optimum values After you ve found an appropriate area on the slide click the Measure Intensity tab in Calibrate Scanner Choose the Filter Set for the wavelength you want to measure GppliedPrecision Chapter 11 Maintaining the System 135 Cleaning Slides To acquire optimal images it is a good idea to clean slides thoroughly before placing them in the scanner Dust or other artifacts can affect image quality and should be removed as thoro
61. canning a slide Figure 30 Area Tab Area Channels Image Advanced Origin x Y B152 160 Width Height 22 40 21 59 Area units are millimeters Origin X Y The distance in millimeters of the bottom left corner of the Scan Area rectangle from the bottom left corner of the slide oriented as it is represented in Slide View Width Height The width and height of the scan area in millimeters Note that width is the long side of the slide height is the short side of the slide Figure 31 Channels Tab Area Channels Image Advanced No Channels fa Filter Exposure 16 Eys x oso Active Channel 1 F Wheel Default No Channels The number of channels to be scanned Filter Determines the filter set to be used for each channel The filter set determines the dye or wavelength that will be acquired Exposure The number of seconds that the slide will be exposed to light while each channel is being scanned Active Channel The channel that will be used for a preview scan For example in Figure 31 the active channel is Channel 1 which is Cy3 See Also Use Preview to Define the Scan Area 34 04 720046 000 Rev D 0804 82 arrayWoRx Standard amp Auto User s Manual Patent Pending Figure 32 Image Tab Areal Channels Image Advanced Sensitivity High S R Resolution a252 OOo Resolution Calculator stripe Scan l Deconvalve _ Resolution units are microns S
62. ckly release it Using the middle mouse button click in a partially hidden window to make the window active means to point to any place in the visible part of the window press the middle mouse button and quickly release it Double click Press the left mouse button and release it two times in quick succession For example double click the arrayWoRx icon to start the Scan software means to point to the icon and press and release the left mouse button two times quickly Right click Press the right mouse button and quickly release it To right click an object on the screen point to it and then press the right mouse button and quickly release it Drag Move an object on the screen to a new position by pointing to the object holding down the left mouse button while moving the mouse pointer and releasing the button when the pointer is at the new position ppliedPrecision Chapter 1 Getting Help Using Online Help The online Help system uses the workstation s Web browser to display its contents Note You can view the Help without an Internet connection because all of the Help files reside on the workstation To open the Help window In the Main Scan window Help menu choose OVERVIEW OF SCANNING to open the Help window array WoRx e Scan File Scan Utilities Help Overview of Scanning 8 B scanner Information Slide View software Versions Or While using a feature with a Help butto
63. cted 04 720046 000 Rev D 0804 38 arrayWoRx Standard amp Auto User s Manual Define the Advanced Options The Advanced tab allows you to specify that bar code data be used for the file name automatically generate images in TIFF format set the gain and enable stitch panel flattening and connection By default all options on the Advanced tab are inaccessible grayed out to prevent accidental changes These options are typically not changed after they are set To enable access to options on the Advanced tab 1 On the Main Scan Window choose File PREFERENCES to open the Preferences dialog Show ToolTips for Icons ii Show Toolbar Icon Text F Enable Advanced Mode M Done Help 2 Click Enable Advanced Mode 3 Click Done to close the Preferences dialog Advanced Tab Options Figure 12 The Advanced Tab Area Channels Image Advanced Use Barcode For File Names Enable Stitch Panel Flattening F Enable Stitch Panel Connection Gain for a Generate Images in TIFF Format J Flip _J Flip Rotate J L Before After See Also Exposure Resolution and Sensitivity 51 GppliedPrecision Chapter 4 Scanning Slides 39 Use Barcode For File Names The Scan Table includes a Data Name column that displays the slide identifier To use bar code data for the slide identifier displayed in this column enable this option Note When this option is enabled all files related to the scanned slide e g
64. e Image tab in the Main Scan window Select a pixel size in Resolution Area Channels Image Advanced Sensitivity High SNR Select a pixel size Resolution 13 0080 15 0000 16 2600 19 5120 Note Pixel sizes that appear with an asterisk in the drop down box are interpolated using the next lower multiple resolution of the base e g selecting a resolution 10 000 will interpolate from 9 756 When you select a pixel size the approximate amount of memory required to store all of the files associated with the scanned image appears in the Status box The file set size is determined by the resolution and size of the scan area as well as the number of channels and whether TIFF files are created Select the Sensitivity In most cases use the High SNR sensitivity setting For unusually bright samples you may need to change the sensitivity setting to High Dynamic Range If you are performing a single channel scan with an exposure time greater than 15 second select Stripe Scan mode For shorter exposure times unselect Stripe Scan Note Stripe Scan enables a fast scanning mode in which the shutter is left open and the exposure time is controlled by the speed of the stage motion If you specify an exposure time that is shorter than the minimum scan time the minimum time is used Stripe Scan is automatically disabled for presets that contain more than one channel Panel Scanning mode is used when Stripe Scan is not sele
65. e Scan Table can include up to 50 entries i Note additional entries in Scan Table after Slot 1 has been selected for Multi Scan Different Presets can be selected for each multi scan pARRRRREED Disable Deselects the Scan toggle buttons on all of the slides in the table so that none of the slides are scanned 04 720046 000 Rev D 0804 34 arrayWoRx Standard amp Auto User s Manual Reset Applies the selected preset to all of the slides in the table This option also enables single scans for each loaded slot removes all multi scans and resets all slide data names Notes 1 The Multi Scan dialog is displayed only for slides that are detected by the system 1 e slides that appear in the Scan Table 2 When you multi scan a slide with Use Barcode For File Names selected array WoRx creates a file name for each scan by appending a scan number to the barcode 3 When more than one slide is selected the Preview area is cleared between scans When performing a multi scan on a single slide with no other slides selected the Preview area is not cleared between scans Cassette Idle Mode To protect the interior of the scanner from dust and other debris an open cassette door closes after a period of scanner inactivity and the scanner enters Cassette Idle Mode This mode is triggered when the lamp cooling fan is automatically turned off four minutes after the lamp is shut down either manually or automatically If the
66. e next smallest non interpolated resolution and then processed by the software to generate an image at the interpolated resolution For example an image acquired at a resolution setting of 8 00 is collected at 6 50 The software then interpolates pixel sizes to 8 00 To refine the resolution 1 Define a small test scan area that encloses typical spots and scan the test area 2 Open the file in the Image window if it does not open automatically If necessary zoom in on the spot until it fills most of the Image window 3 Click Tools MEASURE DISTANCES Then choose Microns as the Units Measure the diameter of the spot Click a point on the circumference of the spot and click another point on the circumference directly opposite the first point The line that is displayed should approximate the diameter of the spot 5 Divide the displayed value by ten This is the minimum resolution 1 e biggest pixel that you should select to scan the slide Applied Precision LLC recommends this method Other methods can be used Note You can also use the Resolution Calculator to estimate the pixel size 04 720046 000 Rev D 0804 54 arrayWoRx Standard amp Auto User s Manual Sensitivity The sensitivity determines whether binning is done in hardware or software and the method used to convert the photon count from the CCD camera into an intensity value For most slides use the default sensitivity setting High SNR Change the sensitivity s
67. e username Command interpreter bin bash User ID Linux assigns an unused User ID number to the account As necessary choose options on the Params and Privileges tabs for the user account Click Accept Enter a password for the account in Changing password and confirm it Click Quit to close LinuxConf oo a dae ae Use the following instructions as a guide to change the Sawfish window manager settings for a new user The procedure will vary depending on the version of Sawfish installed on your system B To change settings in the Sawfish window manager A CAUTION You can render your window manager inoperable by changing settings in the Sawfish window manager If you need help making window manager settings please contact the Applied Precision Hotline 1 On the Gnome Main menu choose Programs SETTINGS DESKTOP WINDOW MANAGER to open Control Center 2 Check that Sawfish is the current window manager 3 Click Run Configuration Tool for Sawfish to open Sawfish configurator 4 Select Expert in Show options suitable for __ users 5 Inthe Sawfish tree select Focus 6 Select enter exit in When does the mouse pointer affect the input focus 7 Inthe Sawfish tree select Move Resize 8 Select box in How windows being moved are animated 9 Select box in How windows being resized are animated 10 In the Sawfish tree select Bindings 11 Select Window in Context 12 Select Button1 Click1 13 Click Delete 14
68. ean of a spot s intensity divided by the mean of its local background This ratio commonly and erroneously called signal to noise or SNR is often used as a measure of scanner performance If the same microarray spots are imaged with two or more scanners the instrument that delivers the highest signal to background ratio is usually judged superior Unfortunately using the average spot and background intensities to generate a simple ratio is not an effective gauge of scanner performance The problem is that this ratio doesn t consider signal variation which can have significant effects on the data Since noise is simply another name for variation any calculation of signal to noise by definition must account for variation in the measured data The commonly used signal to background measure doesn t do this which often leads to faulty conclusions 04 720046 000 Rev D 0804 146 arrayWoRx Standard amp Auto User s Manual Example Two Scanners One Spot In Figure 49 two scanners have been used to image the same spot Figure 49 The same spot imaged with two different scanners Scanner A Scanner B The same spot imaged with two different scanners yields significantly different spot and background variation in addition to different average intensities the horizontal lines Although the background of the image from Scanner A averages slightly over 200 counts it has low variability and the spot is readily detected T
69. edPrecision Chapter 2 System Overview 15 Caution and Warning Labels Hazardous Voltage Warning Label WARNING HAZARDOUS VOLTAGE Hazardous Voltage ENCLOSED Enclosed Voltage or current hazard sufficient to cause Voltage or current hazard sufficient shock bu rn or death to cause shock burn or death Disconnect power cord before Disconnect the system by unplugging the power servicing cord before servicing Heavy Objects Caution Label CAUTION HEAVY OBJECT Heavy Object Can cause strain or back injury Can cause muscle strain or Use proper lifting techniques when moving back injury Use proper lifting techniques when moving Equipment marked with this label weighs 25 1b or more and should be lifted by two people Hot Surfaces Caution Label CAUTION HOT SURFACES HOT SURFACES Do not touch Do not touch Allow surfaces to cool prior to servicing Allow surfaces to cool prior to servicing 04 720046 000 Rev D 0804 16 arrayWoRx Standard amp Auto User s Manual ppliedPrecision CONTENTS Turning On the Scanner and Workstation Opening the Main Scan Window Closing the Main Scan Window Closing the Main Scan Window Turning Off the Scanner and Workstation The Scanner Lights STARTING 3 AND SHUTTING DOWN THE SYSTEM This chapter describes how to turn on and turn off the system and start and close the scan software It also describes the scanner lights 04 720046 000 Rev D 0804 1
70. eference 93 View Menu The commands in the View menu allow you to change the appearance of the display in the Image window The commands do not affect the data stored in the image file F Show Min Max Select Image Colors Select Graphics Colors Zoom Up Zoom Down Color Allows you to toggle between color and grayscale display of the image in the Image window When in Color mode up to three channels may be displayed at the same time When in Grayscale mode only one channel may be displayed at a time Tip You can customize grayscale mode to display an inverted grayscale or a variety of color maps To select a color map choose VIEW SELECT IMAGES from the Image window to open the Select Image Colors dialog and click Modify Grayscale Colormap Show Min Max In Grayscale mode allows you to change the color of pixels with intensity values of zero to cyan and saturated pixels with maximum intensity values to magenta Select Image Colors Allows you to choose the colors used to represent each channel The default image colors are green for wave channel 1 and red for wave channel 2 Select Graphics Colors Allows you to choose the colors of lines that are displayed in the Image window by the Line Profile and other tools To change the color of the scale bar choose the Display command in the Options menu see Options Menu page 94 Zoom Up Makes the image in the Image window larger Note You can also use the
71. ence 83 Use Barcode For File Names The Scan Table includes a Data Name column that displays the slide identifier To use bar code data for the slide identifier displayed in this column enable this option Notes 1 When Use Barcode For File Names is enabled all files related to the scanned slide e g panel TIFF etc use the bar code data in their filenames 2 When you multi scan a slide with Use Barcode For File Names selected array WoRx creates a file name for each scan by appending a scan number to the barcode Enable Stitch Panel Flattening Uses multi panel data to refine the Flat field calibration The image must contain a minimum of 16 panels in order to perform panel flattening Enable Stitch Panel Connection Normalizes intensities between the edges of panels Gain Simulates the effects of turning up the gain on a laser PMT scanner Generate Images in TIFF Format Generates TIFF files automatically after the scan The Flip and Rotate options allow the TIFF image data to be rotated around the X and Y axes Panel View Panel View is the window and group of buttons on the first tab in the lower right corner of the Main Scan window that allows you to view a panel in detail Figure 34 Panel View Panel View Scan Table 04 720046 000 Rev D 0804 84 arrayWoRx Standard amp Auto User s Manual Panel View Window Contains the image of an acquired panel The colored bar at the bottom of the window indicates
72. ensitivity Determines the method used by the software to convert intensity data from the CCD camera into image data Resolution The size in microns of the area on the slide represented by a single pixel in the image Resolution Calculator Opens a tool to help you decide what resolution to use for a scan Stripe Scan Enables a fast scanning mode in which the shutter is left open and the exposure time is controlled by the speed of the stage motion If you specify an exposure time that is shorter than the minimum scan time the minimum time 1s used The specified and actual exposure times are recorded in a log file Stripe Scan is automatically disabled for presets that contain more than one channel It is off by default Deconvolve A method designed to remove the small amount of blurring that is present in images obtained by all optical instruments Applied Precision s proprietary deconvolution algorithms preserve the amount of light throughout the image Blurred data is reassigned to its original location Use deconvolution to eliminate bright spot blooming Select this option when array spot sizes are bright or very large Figure 33 Advanced Tab Area Channels Image Advanced J Use Barcode For File Names Enable Stitch Panel Flattening PT Enable Stitch Panel Connection Gain or I Generate Images in TIFF Format Flip IFlip x _J Rotate C d C d Before After GppliedPrecision Chapter 7 Main Scan Window Refer
73. er in which scanned files are stored click Choose Data Folder and browse to a directory Directory path lengths up to 177 characters are supported If you enter a directory path that exceeds that length the path name is truncated 3 To change the default output file name enter a name in the Job Name field File name lengths up to 70 characters are supported 4 Ifyou are using arrayWoRx Auto and a slide is loaded you are prompted to choose one of the following options e Choose Current Slide Preset to scan only the slide that 1s loaded using the preset that is currently selected for that slide in the Scan table The file name of the scanned image is automatically created from the job name Or e Choose Scan table to scan all of the slides that are selected in the Scan table using the preset settings specified in the table The name of the directory in which the scanned images are saved is automatically created from the job name While arrayWoRx is scanning the file name of the current scan is displayed above the Slide View window When arrayWoRx is not scanning the number of the loaded slide is displayed The Status bar indicates the progress of the entire scan job The relative intensity of each panel appears at the bottom of the Panel View 04 720046 000 Rev D 0804 42 arrayWoRx Standard amp Auto User s Manual Start Scan button Figure 14 The Scan Button and Status Bar EFFET Number of loaded slide Min Ma
74. etting if there are dim spots with biologically meaningful information 1 e spots are dim because of under regulated gene expression not because of dye degradation or other slide quality reasons or there is a wide distribution of spot intensities on the slide Table 6 Sensitivity Settings Sensitivity Setting When to use it High SNR Default For typical slides Hardware binning 14 bit camera signal scaled to 16 bits Maximum signal to noise ratio High Dyn Range When there are both very dim and very bright spots Software binning SE aay 14 bit camera signal scaled to 16 bits Binned pixels averaged to minimize saturation If bright spots are saturated and decreasing the exposure time does not eliminate saturation try changing the sensitivity setting 1 Perform a preview scan with the CY3 channel 2 In Slide View find a bright spot that appears saturated on the display 3 Drag the panel tool to the spot 4 Look at the value displayed in Panel View to confirm that the data are saturated Saturation occurs at 125 of optimum GN Adjust the exposure until the Percentage of Optimum is near 100 6 Ifthe brightest spots still saturate with the shortest exposure time try using higher resolution images smaller pixel size to reduce the image sensitivity The shortest exposure 1s from 0 07 to 0 10 depending on the system s shutter 7 Repeat Steps 3 and 4 to verify that the spot is no longer saturated 8 Ifthe spo
75. f intensity values 96 Delete file 112 Preset 49 user account 122 Desktop Gnome 110 starting scan from 19 DHCP 12 Dim image correcting 137 Dim spots changing sensitivity for 54 correcting exposure time 137 Dimensions measuring 69 slide and microarray 57 Directory shared data 12 Disable button 33 85 Disk mount and unmount 113 Display intensity 68 setting attributes 94 wavelengths 88 Distance measuring 69 Document conventions 3 Double click 4 Downloading log files 77 Drag 4 Drive mount and unmount 113 Dyes choosing filter set for 36 supported 144 Dynamic range and sensitivity 37 Percent of Optimum 55 sensitivity option 54 setting 37 true dynamic range 147 using High Dyn Range 54 E Eject cassette 27 44 slide 77 Eject button 78 Electrical requirements 11 Electrostatic discharge 129 Emission filter 9 Enable Stitch Panel Connection 83 Enable Stitch Panel Flattening 83 Environmental requirements 11 Excitation filter 9 Excluding scan areas 43 Exit Main Scan window 21 Exporting TIFF files 102 Exposure and intensity 52 55 defined 36 52 for preview scan 48 ppliedPrecision guidelines 52 55 ideal 55 setting 81 typical settings 52 Extending scan area extending 58 F Fast scanning mode 37 82 Feature Size 56 defined 56 File choosing data folder for scan results 48 copy 112 delete 112 find in File Manager 112 move 112
76. face Legend Allows you to show or hide a key showing color and value equivalents Keep Scale Constant When Keep Scale Constant is selected the minimum and maximum intensity value of the entire image defines the range of the graph Otherwise the graph is scaled using the minimum and maximum values of the region of interest Note When both Keep Scale Constant and Legend are on all cells in the legend may not be visible Add Optional Title Creates and displays a title for the graph 04 720046 000 Rev D 0804 100 arrayWoRx Standard amp Auto User s Manual Histogram The Data Inspector Histogram displays the intensity distribution of a region of interest in the Image window More detailed information about using the Histogram is available in online Help Click Help in Data Inspector Histogram or Histogram Calculation Options to display help for those tools Figure 40 Data Inspector Histogram Data Inspector Histogram Graph Properties Options Print Help Close Graph Properties Allows you to change the appearance of the histogram Options Allows you to change the parameters of the histogram GppliedPrecision CONTENTS Overview Converting an Image File to TIFF Saving Files to CD 9 SAVING IMAGE DATA Overview The arrayWoRx software creates and saves image files for each slide that is scanned These files are saved in an Applied Precision proprietary format that has a
77. front bail is positioned over the slides so that the slides do not slip out of the cassette 3 Press the Load Eject button to open the cassette door 4 Place the back arrow end of the cassette into the scanner at an angle by gently setting the bottom back end of the cassette down in the back of the cassette tray ppliedPrecision Chapter 4 Scanning Slides 31 5 Lower the front end of the cassette so that its guide slots slide over the green guide pins on the inside front of the cassette tray Once the pins are set fully into the slots the guide pins will hold the cassette in place Fa ln Guide Pins 6 Press the Load Eject button The scanner will pull the cassette into the system close the door check to ensure proper cassette positioning and inventory the slides If the cassette is not positioned properly the system will automatically reopen the cassette door eject the cassette and display an error message If this occurs follow the instructions that are displayed in the error message The Status light will shine steady amber once the cassette is loaded After the scanner removes a slide from the cassette and loads it for scanning the Status light will shine a steady green Viewing Slide Status Once a cassette has been loaded and inventoried all slots with detected slides will be displayed on the Scan Table tab of the Main Scan dialog All of the detected slides will be automatically selected for
78. g down Insert opaque slides with arrays face up ie 3 Repeat Step 2 for each of the slides that you want to insert into the cassette 4 After all the slides have been loaded into the cassette move the front bail so that it is positioned over the back edge of the loaded slides AN CAUTION When you are loading slides do not place the cassette with the rear bails on a flat surface such as a table or counter top This can open the rear bails and cause the slides to fall out of the cassette when it is moved 04 720046 000 Rev D 0804 30 arrayWoRx Standard amp Auto User s Manual Load the Cassette into the Scanner An arrow on the top of the cassette points to the back of the cassette The back of the cassette also has a double rather than single bail When loading the cassette into the scanner the back of the cassette is inserted first To load a cassette into the arrayWoRx Auto scanner 1 Check the Status light e Ifthe Status light is shining a steady green or amber there is already a cassette in the scanner Click the Eject button on the Main Scan window or press the Load Eject button located in front of the cassette door to eject the cassette Refer to the figures in the previous section or e Ifthe Status light is off and the cassette door is closed the scanner may have entered Cassette Idle Mode page 34 To exit from Cassette Idle mode press the Load Eject button on the scanner 2 Ensure that the
79. he slides in the table Then select the Scan column toggle buttons for the slides that you want to scan Load Unload a Single Slide To load a single slide 1 Click the Load Slide option to display the list of slides 2 Click the slide to load The slide will be removed from the cassette and positioned for scanning The scanner Status light flashes and turns green when the slide is ready ppliedPrecision Chapter 4 Scanning Slides 41 To unload a single slide 1 Click the Load Slide option to display the list of slides 2 Click None The loaded slide will be returned to the cassette Note In general it should not be necessary to unload a slide When you click the Load Slide option the current slide is automatically unloaded before the next slide is loaded Likewise when you have finished viewing the contents of a cassette the cassette should be ejected This unloads any currently loaded slide Start and Stop the Scan Before you start the scan make sure that the settings on the Area Channels Image and Advanced tabs are correct Note If arrayWoRx displays a message that indicates the camera temperature is too high wait until the message closes before you try to perform a normal scan To start the scan 1 Click the Scan button in the Main Scan window to start the scan The Scan Options dialog opens and displays the default output Data Folder and the Job Name 2 To change the output fold
80. he GIMP to View TIFF Files ccccccccccccceceeeeeeeees Capturing a Screen or Window in Gnome cccceeeeeees Editing Files with the gedit Text Editor cccceeeeeees Managing User ACCOUNHS ccccccccccccecccccceeeeceeeeeeceeeceeeeeeeees Usine Wai COME oiean ira EONA raai Changing a User s Password ccccccceeeeseeeeeeeeeeeeeeeeeeeeeeeeeees Delete a User Acconto aaa Creating a New User Account seesseeeseseeeerrererrerrrreeee Fle Permissions e a a Changing the Workstation Time Date nnnnnnennneeneeeeneeeen Guidelines for using LinuX ssessssessssesseesseseseseeeerrrrerreeeeeeeee 11 Maintaining the System We aS WhO TLC SHY renien ia N E E Cle aint SSCS anche asa tek A 12 Troubleshooting Problems with Tag CS osoro srie a Oher PEODICINS nieta n a E tessices Other Problems Contd eue cenarireaneni e a i Running the Service Script cccccccccccccecccccecececcseceeeeeseeeseees arrayWoRx Problem Report Form cccccccceeeeeeeeeeeeeees Appendix A Filters and Dyes Filters and Dyes supported by arrayvWORKX ccccccccecssseeeeseees Appendix B Signal to Noise Contents V Index 04 720046 000 Rev D 0804 vi arrayWoRx Standard amp Auto User s Manual ppliedPrecision CONTENTS Using This Book Document Conventions Using Online Help Contacting Applied Precision LLC 1 GETTING HELP The arrayWoRx documentation consists of
81. he ROI If Ratio is selected the table displays the ratio of intensities of two channels for each pixel for example Cy3 Cy5S File Menu Allows you to save the currently viewed table of data in an ASCII text file for off line processing open a file to collect statistics records or close the Data Inspector View Menu Allows you to create a 3D graph or a Histogram display Options Menu Allows you to choose options for the format of the Statistics Record output file select options for subtracting background intensity from calculations choose the wavelengths to use 1n ratio calculations and modify settings number of bins bin width etc for histogram plots ppliedPrecision Chapter 8 Image Window Reference 97 Current Window The window number of the Image window from which data is being displayed in Data Inspector Select Box Allows you to define a region of interest ROI in the Image window Click Select Box and then move the cursor to the Image window and select a region to define the ROI The data from this ROI is then displayed by Data Inspector whether in the Data Table Histogram or 3D Data Graph Columns Rows The size of the region of interest Columns is the number of horizontal pixels and Rows is the number of vertical pixels in the region of interest You can define the size by entering the values in the fields or by clicking Select Box and defining the ROI graphically Constrain to Circle When Constrai
82. he image from Scanner B has significantly more noise 1 e variation in both the spot and background intensities Consequently the spot imaged by Scanner B cannot be detected using standard criteria This difference is correctly interpreted as better performance by Scanner A due to its superior ability to separate the spot s signal from the local background when including effects of variability The intensity plots shown in the lower panels indicate that Scanner A has a less desirable signal to background ratio than Scanner B In particular the average background intensity of the image from Scanner A is about twice that of Scanner B significantly reducing its signal to background ratio With greater separation of the spot s average intensity from its average local background Scanner B would appear to provide better detection Clearly however this is not the case The spot imaged with Scanner A can be seen quite easily whereas the same spot imaged with Scanner B is much more difficult to distinguish Why does the signal to background metric fail to identify the clearer of the two spot images Because it ignores variation in the spot and background measurements When this is taken into account the image from Scanner A is correctly identified as providing superior detection of the spot GppliedPrecision Appendix B Signal To Noise 147 Image Noise All microarray images have noise or variation in the imaged signals Noise comes from t
83. he import command from the GNOME Help index click Man Pages and click import in the list of user commands To capture a screen or window 1 Ifthe Screenshooter icon is not on your Gnome Panel click the Main Menu button ba e in the Gnome panel and select Applets UTILITY SCREENSHOOTER 2 Capture a screen window or area as follows e To capture a screen shot of the entire desktop click Or e To capture an image of a specific window or area click and click on the window or drag the cursor across the area that you want to capture Note You can also use the GIMP see page 115 to acquire screenshots by choosing File SCREENSHOT ACQUIRE SCREENSHOT In the GIMP Main window You can choose to acquire an image of the entire desktop display or to acquire an image of a single window with or without decorations overlays 04 720046 000 Rev D 0804 118 arrayWoRx Standard amp Auto User s Manual Editing Files with the gedit Text Editor You may occasionally need to edit text files This topic describes how to open a file with a text editor and how to save and close the file If you need more detailed information about using gedit you can download an HTML User s Manual at http gedit sourceforge net To open a file in gedit 1 Choose PROGRAMS APPLICATIONS GEDIT from the Gnome Main menu 2 Choose OPEN from the File menu and select the file from the Open File window Note You can also open a file by d
84. ic IP address using Windows Network and Dialup Settings Private Virtual is the connection to the shared data directory in Linux It is configured to use a static IP address of 172 16 197 2 There is no physical connection for this communication path it is accomplished via Samba in Linux and default virtual Ethernet port settings in VMWare CAUTION Private Virtual settings should NOT be modified Modifying this setting causes communication with the shared data directory to fail Do not change the Windows 2000 account awuser or its password Changing this account name or password will cause the workstation to become non functional due to internal networking conflicts Cable Connections The scanner and workstation are connected by a single ethernet cable Note that the workstation has two ethernet ports one for the scanner and the other for a network connection Figure 3 shows the correct cable connections Figure 3 Scanner and Typical Workstation Cable Connections Workstation UT Mouse oar BS diassan jann seen susesusuenes ah rT ee ef ee oe EEH ETT fit ce aar fi lt Ethernet to we Scanner Iar TE Monitor A er DOD Ethernet to Ethernet to a workstation network Note The exact location of connectors on the workstation will depend on the computer included as part of the system 04 720046 000 Rev D 0804 14 arrayWoRx Standard amp Auto User s Manual Safety The arrayWoRx sys
85. ile menu commands allow you to open image files save them in different formats and print them Image Window 1 mageFile2 stc File View Options Tools Ctrl S Save As TIFF Ctrl T Print Get Info Ctrl I Close Window Open Allows you to choose an image file to open in the Image window Save Saves the image or a portion of the image in the Image window to a file You can choose how intensity values are scaled in the saved file Save As TIFF Saves the image or a portion of the image in the Image window to a TIFF file You can set scaling parameters for each channel and save each channel to a separate TIFF file See Converting an Image File to TIFF on page 102 Print Prints the image in the Image window to a printer or file A printer must be connected to the workstation and Linux must be configured properly If you need to print to a printer on a Linux or UNIX network contact Applied Precision Customer Service Note Another way to print images is to save them in TIFF format in the shared_data drive open Microsoft Windows through VMWare and print the TIFF files to a printer on the Windows network Get Info Displays information about the image in the Image window such as minimum maximum and mean intensity values size of the intensity table and the type of lens used to acquire the image Close Window Closes the Image window GppliedPrecision Chapter 8 Image Window R
86. ion 39 Stitch panel flattening 39 Stop a scan 41 Stop button 78 Stripe Scan 82 when to use 37 Switch operating systems between Linux and Windows 18 while shutting down 19 T Task buttons Gnome 110 Technical support 6 11 Temperature operating 11 04 720046 000 Rev D 0804 Index 155 Text editor 118 TIFF converting image file to 102 format options 102 generate automatically 39 opening 67 rotating 83 rotating image 39 save as 92 TIFF Viewer See GIMP Time estimated scan time 86 guidelines for exposure time 52 minimizing scan time 52 Tool tips turn on or off 74 Toolbar show or hide 94 Tools Data Inspector 95 hide or show 94 Image window 66 Line Profile 95 Measure Distances 95 set display attributes 93 Tools menu 95 Turn off system 18 Turn on system 18 U Ultraviolet light 14 Unmount drives 113 URL Applied Precision 6 Use Barcode for File Names option 34 User accounts creating 123 managing 119 User Mount 113 Username first time user 18 User s Guide Gnome 109 Utilities menu Main Scan window 76 V Version software 76 77 View channel intensities 68 image files 67 TIFF files 67 View menu 93 Viewing scan data 44 VMWare 10 W Warning 156 arrayWoRx Standard amp Auto User s Manual defined 3 Warning labels 14 Wavelength selector 68 88 Wavelengths display intensity of 68 measuring intensity 134 select colors for 93 selecting 68 sho
87. k OK again 7 Restart the system to verify proper functionality Note Clicking Cancel on the Restoring connection to message box disables the shared data directory link between Windows and Linux for the session Running the Service Script Follow these instructions if an Applied Precision technician asks you to provide system diagnostic files To run the Awe Service script 1 Turn on the workstation and scanner log in as awuser and open the ArrayWoRx software Double click the Home Directory icon on the desktop 3 Make sure that the scanner is on The scanner is on if the Power LED 1s green or flashing green or if all three LEDs are flashing amber 4 Browse to the usr local bin directory and double click the service term sh file If your system doesn t have this file contact APLLC to get installation media and follow the instructions in To install the Awe Service script on page 140 5 Follow the directions in the terminal window The Scanner Service files are collected and placed in the service tar z file in the tmp service directory This file is compressed and ready to send to APLLC 04 720046 000 Rev D 0804 140 arrayWoRx Standard amp Auto User s Manual To run the Awe Service script with verbose output l 10 Turn on the workstation and scanner log in as awuser and open the ArrayWoRx software Double click the Home Directory icon on the desktop Browse to the usr local bin direc
88. k the Gnome Main Menu button Gnome Main we Menu Button A e E 5 5 Choose LOG OUT on the Gnome Main menu Programs he Favorites Applets gr KDE menus BF Run oy Panel 2a Lock screen J Click Logout 8 tos out YO Yon See Also Working with Windows 111 ppliedPrecision Chapter 3 Starting and Shutting Down the System 23 6 Choose Halt on the Logout dialog and then click Yes Gnome and Linux will close and the workstation will turn off P Really bog oul J Save current setup Lgi Choose Halt Jana Click Yes gt gt w _ mw 7 Press the Power button on the front of the scanner to turn off the scanner The Power light flashes while the system shuts down and turns off when the system is completely shut down 8 Ifthe Power light stays on and steady amber for more than 20 seconds press the Power button a second time to turn off the system Note Before the scanner completely shuts down the Power light turns steady amber 9 Turn off the monitor The Scanner Lights The Power Status and Lamp lights on the scanner provide information about the status of the scanner Each light has five states green steady green flashing amber steady amber flashing and off Table 3 shows the possible states of each light and what each state means Scanner Lights 04 720046 000 Rev D 0804 24 arrayWoRx Standard amp Auto User s Manual Table 3 Scanner Light States and What
89. lamp assembly provides light to the optical system The assembly consists of a metal halide bulb and lamp housing After 1000 hours of use the bulb in the lamp assembly can no longer supply the intensity and spectral range of light required by the scanner s optics The scanner software keeps track of the hours of use for the bulb and after 1000 hours the Lamp light on the front of the scanner changes to amber In order to ensure that the bulb is properly aligned in the optical system replace the entire assembly as a unit Note Use only replacement bulbs supplied by Applied Precision Safety There are three risks associated with handling the lamp assembly exposure to heat exposure to ultraviolet light and exposure to mercury vapor Oil from fingerprints on the bulb can heat up and cause the bulb to explode when it is turned on releasing dangerous vapor Observe the following safety precautions when handling the bulb assembly e Do not open the lamp access door while the lamp is turned on e Wait for the lamp assembly to cool before handling the assembly e Do not touch the bulb Use lens tissue or a lint free cloth to handle the bulb Clean fingerprints from the bulb using an organic solvent such as ethanol and a lint free cloth Note The lamp access door is equipped with a safety interlock If you open the lamp access door while the lamp is turned on the safety interlock turns off the lamp WARNING Under certain conditions
90. le20 config _xx tif A A E EEU IEEE EE i D a z5canFiles i Scale2005000_w6 Scale 85 tif l out_files released_help 125 267 476 bytes in 1 file ppliedPrecision Chapter 9 Saving Image Data 105 4 Drag each file that you want to copy from the File Manager to the CD Archive Manager file window As you put files in the file window the status box directly above the window tells you how much space the files require and the percent of space remaining on the CD Tip You can select more than one file in the File Manager by holding down the CTRL key while selecting files 5 When you have placed all of the files that you want to copy into the file window click Create CD to open Create CD Create CD F Add CD Catalog Entry fT Save CD Catalog On Disc Catalog Entry Name AWVolume Catalog Note Archiver Name awuser Create CD Cancel 6 Ifyou want to keep a list of CDs that have been created using Archive Manager select Add CD Catalog Entry and or Save CD Catalog On Disc Type a name note and archiver name to be recorded in the catalog in the boxes as needed Add CD Catalog Entry adds this entry to the local catalog stored on the workstation Save CD Catalog on Disc saves the catalog on the newly created CD Click Help for more information about the catalog 7 Click Create CD to start copying files to the CD Wait for the message CD Creation Complete to appear in the Archive Manage
91. light and heat hazards 14 Sampling 56 Saturated spots 55 Saturation 52 Save for Linux and Windows 12 in shared data directory 12 Save as TIFF 92 files 88 files in gedit 118 files to CD 74 for Linux and Windows 101 images 92 intensity data to ASCH files 96 preset 80 rename preset 80 select region to TIFF 103 user account data 123 Save Scan settings 48 Sawfish window manager 111 Scale adjusting intensity scale 91 enabling the Scale bar 89 intensity setting 68 Scale Bar 89 Scale Image 90 Scan at low resolution 75 canceling 75 displaying estimated time 86 file size 86 finding boundaries 35 minimizing scan time 52 multiple scans of a slide 33 normal 75 preview 75 progress indicator 86 width and height 81 Scan area 34 cropping 35 moving 34 rectangle 79 resizing 35 selecting 34 Scan Area extending 58 Scan button 32 78 Scan menu Main Scan window 75 Scan Options 48 Scan Settings saving 48 using presets 80 Scan table 31 84 Scan starting and stopping 41 Scanner applications 7 components 8 evaluating performance 148 focusing 60 Scanner disk usage 77 Scanner Lamp replacing lamp assembly 130 turning on or off 76 Scanner memory 77 Scanner Software Package Status 76 Scanner connecting to workstation 13 Scanning an area with different settings 43 excluding areas 43 Screen capture 117 Screenshooter 117 Scroll Bars 89 SELECT Box IN DATA INSPECTOR 72
92. ll files related to the scanned slides e g panel TIFF etc will share the name displayed in this field Note Data name lengths up to 70 characters are supported ppliedPrecision Chapter 4 Scanning Slides 33 Load Slide Displays a list of all slots available in a cassette Individual slides may be loaded and unloaded by selecting them from the list Slots containing detected slides are displayed without parentheses empty slots are displayed with parentheses Tip You can use this option to try manually loading a slide that does not appear in the Scan Table but is known to be in the cassette On rare occasions a slide is not detected during inventory If a slot contains a slide but it appears in the list with parentheses you can select that slot and the scanner will attempt to re detect the slide If the slide can be successfully detected and loaded it will appear in the Scan Table however parentheses will remain around the slide in the Load Slide list Multi Scan Allows a single slide to be scanned multiple times If only one slide is present it is automatically added to the Scan table If more than one slide is present a dialog opens that allows you to choose which slide to multi scan Once a slide is selected it appears on a second and if applicable third fourth etc line in the Scan Table This allows the same slide to be scanned with different presets exposure times pixel sizes resolution etc Th
93. loses the preview image of the array If you are defining the scan area for a batch of slides leave some space between the blue rectangle and the array to account for variability in array printing 5 Make sure that the desired channel is marked as active in the Channels tab Notes 1 Choose Scan CLEAR PREVIEW to clear the preview image in Slide View 2 If you know the dimensions for your scan you can enter them manually in the text boxes on the Scan Area tab 04 720046 000 Rev D 0804 36 arrayWoRx Standard amp Auto User s Manual Choose the Filter and Exposure Typical Settings Number of channels 2 Ch 1 Filter Cy3 Exposure 0 10 Ch 2 Filter Cy5 Exposure 0 30 You can choose the filter and an exposure for up to four channels The number of channels that you choose for a scan 1s typically the same as the number of fluorophores that you want to measure Note If you would like to add third party filters to your arrayWoRx scanner contact Applied Precision Customer Service at 1 800 862 5166 or servicehotline api com Channel Each wavelength dye measured during a scan is represented as a channel in the scan software A Filter name and exposure time is associated with each channel Filter The Filter setting 1s named for the dye fluorophore to be measured The scanner uses this setting to determine which filters to use for the scan Exposure The time in seconds that the sample is exposed to light from the lamp
94. lp system The arrayWoRx workstation also uses Windows 2000 however since most users are familiar with Windows operating systems we do not describe how to use Windows in this manual Overview Linux is one of the operating systems on the arrayWoRx workstation Gnome is a user friendly windows based interface between you and Linux Most of the usability of Gnome is similar to MacOS and Windows Although most of the work you do on the system can be done using Gnome you may want to use Linux commands for some tasks To access the Linux command line open the Gnome Terminal Emulator by clicking on the Gnome Panel 04 720046 000 Rev D 0804 108 arrayWoRx Standard amp Auto User s Manual Logging On to and Off of Gnome Gnome Main Menu Click Log out To use the workstation you must first log on to Gnome If you are logging on to a new system for the first time see Turning On the Scanner and Workstation on page 18 for the username and password to use To log on to Gnome 1 Turn on the workstation and monitor 2 When the Login screen appears move the mouse pointer to the Welcome dialog 3 Type the username awuser and press ENTER If a Linux password has been assigned to the system type your password into the appropriate field and press ENTER again Linux and Gnome will start up To log off of Gnome 1 Close all windows and applications on the Gnome desktop Note To close arrayWoRx software choose QUIT or CLO
95. lt gt S amp e It s best to choose letters upper or lower case numbers 0 9 or _ or for filenames The character is also fine e Spaces are not allowed in names e Try to create file names that mean something and can be recognized later e You can have a directory and a file named the same e DeltaVision files have the extension dv OTF files are otf etc Do not change or eliminate filename extensions Memory Buffering Linux caching of memory to the hard drive is very aggressive In particular Linux may pause or slow down more frequently depending on the degree of disk caching To monitor memory buffering e From the menu in the task bar choose Applets Monitors CPU MEM usage to display the memory usage on the task bar Then watch what happens to the bottom bar in the applet the top two are for processor and memory respectively This should give you an idea of swap memory usage if the middle bar goes up to its maximum you ll see the bottom bar start to be used This represents cached memory which is considerably slower System Lockup To escape from a system lockup try simultaneously pressing the C l Alt BackSpace keys which should take you back to the System Log In Screen 04 720046 000 Rev D 0804 128 arrayWoRx Standard amp Auto User s Manual ppliedPrecision CONTENTS Electrostatic Discharge ESD Cleaning the System Replacing the Lamp Assembly Measu
96. may result in loading problems that require out of warranty service repair e Slides with sticky edges should not be loaded in the scanner If slide edges are adhesive enough to pick up a sheet of paper they should be cleaned prior to loading Slides that are loaded in the scanner must have the dimensions shown in Figure 18 Figure 18 Slide Dimensions and Range of Acceptable Array Dimensions F gt H C _ gt Dimension Acceptable Range mm A Slide length 75 0 76 0 B Slide width 25 0 26 0 C Scan area length maximum 60 6 D Scan area width maximum 272 E Bar code length maximum 7I F Barcode width maximum 15 G Slide thickness 0 96 1 04 If your system has the required hardware you can increase the scan area length to 73 6 mm by extending the scan area see page 58 The Length Width A to B angle must be 90 2 04 720046 000 Rev D 0804 58 arrayWoRx Standard amp Auto User s Manual Extending the Scan Area If your system has extended scan area hardware you can extend the scan area to cover the entire slide Note This option is available only for systems that have extended scan area hardware To determine whether your system has this hardware check the serial number Upgraded systems have serial numbers E6XXXX ESA or A6XXXX ESA New systems have serial numbers E65XXX 2 color E64XXX or A64XXX 4 color Figure 19 Extended Scan Area SPHERE To extend the scan area
97. me Restores the display of the image in the Panel view to its original state The image is resized to fit in the Panel view with the image centered in the view The intensities are automatically scaled and restored to their original values TIP The Home button is useful when you want to center the image after you have used the scroll bars Scan Table Scan Table is the second tab in the lower right corner of the Main Scan window It contains the Scan Table along with several buttons that you can use to set up the table The table varies slightly for the arrayWoRx Standard and arrayWoRx Auto products In addition to the Multi Scan Disable and Reset buttons on the arrayWoRx Standard Scan Table the arrayWoRx Auto Scan Table contains the Load Slide option and displays information about the slides and the cassette as shown on Figure 35 Scan Table ppliedPrecision Chapter 7 Main Scan Window Reference 85 Figure 35 Scan Table ae fotz m so 4 ots 4 ots 4 pot 4 pots a pots a Potio 4 Note The arrayWoRx Auto Scan table displays the loaded slots at the top of the table For example if slots 2 4 and 6 are the only loaded slots in a cassette the table displays slots 2 4 and 6 followed by slots 1 3 5 and 7 Load Slide Allows individual slides to be loaded and scanned Slots containing detected slides are displayed in the Load Slide list without parentheses Empty slots are displayed with parentheses See Viewing
98. n click the Help button to display help for the feature To navigate the Help system Choose an item in the Contents pane of the Help window to view the topic Click the SEARCH tab to display the Search pane Type a word or short phrase in the field and click Search or press ENTER to display a list of topics that contain the word or phrase To view a topic click an item in the list of topics Click the INDEX tab to display the Index pane Click on a letter at the top of the pane or use the scroll bar to find an indexed word or phrase To view the topic click the indexed word or phrase Click See Also in the lower right corner of some topics to display a list of related topics Click File CLOSE to close the Help window 04 720046 000 Rev D 0804 5 arrayWoRx Standard amp Auto User s Manual Contacting Applied Precision LLC If you have questions about arrayWoRx first refer to this manual or consult the online Help system If you don t find the information you need contact us at one of the following phone numbers or addresses Customer Service Hotline Phone 1 800 862 5166 email servicehotline api com Hours 8 00AM 5 00PM Pacific Time Monday Friday Corporate Offices Internet Address www appliedprecision com Corporate Headquarters European Office Applied Precision LLC Applied Precision LLC 1040 12 Avenue NW 121 High Street Issaquah WA 98027 Marlborough SN8 1LZ USA United Kingdom Phone 425 5
99. n stc extension You can set scan options to automatically create TIFF files when you save image files or you can convert image files to TIFF files after they are saved After converting image files to TIFF format you can save the TIFF files on a network drive or to a CD Note Save arrayWoRx image files to the default shared_data shared drive on the arrayWoRx workstation Files in this folder can be opened from either Linux or Windows 04 720046 000 Rev D 0804 102 arrayWoRx Standard amp Auto User s Manual Converting an Image File to TIFF This section describes how to convert an image file that is displayed in the Image window to a TIFF file TIFF files can also be generated at scan time using the selections on the Advanced tab Note Before converting a file make sure that a TIFF version of the file has not already been created Look in the data folder that contains the image file TIFF versions of the image file are assigned a name based on the file name with the extension tif You can export files in three TIFF formats 8 bit grayscale 16 bit grayscale and 24 bit RGB color The 8 bit and 16 bit formats create one file for each wavelength The 24 bit RGB format creates one file that includes up to three wavelengths To export an image file to TIFF from the Image window 1 Open the image file in the Image window 2 On the Image window File menu choose SAVE AS TIFF Select Region Reset Details Wavelengths W 5
100. n be viewed in the Image window or converted to the TIFF format to be viewed in a TIFF image viewer 04 720046 000 Rev D 0804 26 arrayWoRx Standard amp Auto User s Manual Handling Slides With arrayWoRx Standard Insert the Slide into the Scanner Insert the slide into the scanner with the array facing down Insert opaque slides face up To be detected bar codes must be on the bottom of the slide facing down Notes 1 Inserting the slide with the array facing down optimizes light collection The refraction of the light as it passes through the slide can improve the signal by a factor of four 2 If the Status light is on and green there 1s already a slide in the scanner If there is a slide in the scanner click the Eject button on the Main Scan window or press the Eject button on the scanner front panel and remove the slide Figure 5 The Eject Button To insert a slide into the scanner e Hold the slide by the end with the label or bar code and gently insert the slide completely into the slide slot Insert the slide with the array facing down unless it is an opaque slide Insert opaque slides with the array facing up The Status light will flash amber as the scanner loads the slide and then turn green when the slide is loaded natan ty PAs AU Jeg y laap gt mn lt yo gt Slide Slot S Lamp Light Sap gt Eject Button When the slide is loaded the scanner s lamp automatic
101. n read from or write to a disk or CD the drive must be mounted to the file system Before you remove the disk from the drive the drive must be unmounted See Mounting and unmounting drives in If you are new to Linux UNIX in the online Gnome User s Guide Note If you look at the contents of a floppy disk or CD in the File Manager and don t see any files the floppy disk drive or CD read write drive may not be mounted 04 720046 000 Rev D 0804 114 arrayWoRx Standard amp Auto User s Manual To mount a drive 1 Insert the disk in the floppy drive or the CD in the CD drive 2 Choose PROGRAMS SYSTEM DISK MANAGEMENT on the Gnome Main menu to open the User Mount tool User Mount Tool Directory Device Type mntfloppy dev fdo auto Mount Format fmntcdrom dev cdrom iso9660 Mount Format 3 Click Mount for the drive you are mounting in the User Mount Tool e mnt floppy to mount a floppy disk e mnt cdrom to mount a CD 4 Click Exit to close the User Mount Tool Before you remove a disk or CD from its drive unmount it If you remove a disk without unmounting the drive data you expect to be written to the disk may not be written Similarly you will not be able to eject a CD from its drive until it has been unmounted You can tell whether a drive is mounted or unmounted by looking at the Mount Unmount button in the User Mount Tool The button will say mount if the drive is not mounted and unmoun
102. n to Circle is turned on Data Inspector only displays intensity data for pixels inside the circle shown by the ROI box in the Image window This makes it possible to display data for the pixels in a spot without including background pixels Show 3D Graph Opens a 3 D graph of ROI pixel intensities in a separate window The graph updates automatically as you move and resize the ROI Show Histogram Displays a histogram of the intensity distribution of pixels in the ROI in a separate window The graph updates automatically as you move and resize the ROI Ratio Changes the values displayed in the table 3D graph and histogram from absolute intensity values to intensity ratios of channels in the ROI Statistics Displays the minimum maximum and mean intensities number of pixels and the standard deviation of intensity values of the image file being displayed in the Image window If Ratio is selected the Statistics field displays the average ratio of intensities in the ROI 04 720046 000 Rev D 0804 98 arrayWoRx Standard amp Auto User s Manual 3D Data Graph The 3D Data Graph displays the intensity or ratio profile of a region of interest in the Image window defined in Data Inspector The graph automatically updates itself when you define a new ROI in the Image window or make other changes in Data Inspector Figure 39 Sample 3D Data Graph 3D Data Graph Sample 3D Intensity Graph J Shaded I Contours IT Zones J Smoo
103. nels that contain bright artifacts Bright artifacts can be caused by preparation factors such as dye clumps dust or the existence of other impurities in the background Generally a slide should be scanned within a few days of being hybridized to avoid the effects of dye degradation For spots that are dim due to dye degradation increasing the exposure time can increase the signal intensity However unless the background degrades along with the spots increasing the exposure time will not improve image quality it will simply make the scan take longer Typical exposure settings are shown in Table 4 Because of the linear response of the CCD camera the intensity is directly proportional to the exposure time For example if an exposure setting of 0 3 second causes the Cy5 channel of the brightest spot to have an intensity of 10 000 counts increasing the exposure time to 0 6 will increase the maximum intensity to 20 000 counts The maximum intensity at saturation 1s 65 536 counts Table 4 Typical Exposure Settings Dye Exposure in Name Seconds Cy3 0 1 0 2 Cy5 0 3 0 5 max 2 sec Tip You can choose exposure settings by examining intensity values in a Preview scan As you move the mouse pointer over the image the colored status bar in the Preview window displays the intensity value of the pixel that is under the mouse pointer Use Preview to Define the Scan Area 34 Choose the Filter and Exposure 36 ppliedPrecision
104. ns For the quickest preview scans the preview exposure time should not be set lower than about 0 15 second this varies slightly for each system If the preview exposure time is set lower than this value the system automatically switches from scanning one column at a time to scanning one panel at a time which is much slower This setting does not affect normal or full scans Working with Presets You can save the options and values that are used repeatedly to scan similar slides in a Preset The Preset defines the following options that are assigned to a scan e Scan Area e Number of Channels e Exposure GppliedPrecision Click Create Click OK to create the set Click Save to overwrite saved settings Chapter 5 Refining Scan Settings 49 e Resolution e Sensitivity After you define presets you can select a preset for each slide in the Scan Table To create a preset 1 Choose the options on the Area Channels Image and Advanced tabs that you want to save 2 Click Create Type a name for the set 3 Type a name for the preset in the Prompt dialog and click OK Note If you change any of the options or values on the Channel Scan Area or Image tabs or move the Scan Area rectangle and click Save the preset is immediately updated and the old settings are erased Be careful not to inadvertently change the scan area or other options in a preset Click Create to save the new preset without erasing the old settings
105. ntensities significantly below 100 of optimum indicate that the dynamic range is not being fully utilized To achieve the best possible signal to noise ratio you may need to increase the exposure time To determine an ideal exposure time for a scan it is best to perform a Preview Scan first to identify the brightest areas of interest on the slide Then using single acquisition and the Percent of Optimum display determine an exposure time that yields non saturating areas while maximizing the signal to noise ratio for the lower intensity areas of interest Using the Resolution Calculator Use the Resolution Calculator to help determine the proper resolution for a scan when you know the approximate size in microns of features on the slide and the level of detail you want in the image 04 720046 000 Rev D 0804 56 arrayWoRx Standard amp Auto User s Manual To open the Resolution Calculator 1 Click the Image tab 2 Click Resolution Calculator Area Channels Image Advanced Sensitivity Resolution 8 2520 Resolution Calculator Resolution Calculator Feature Size microns 130 00 Sampling Optimal Recommended Resolution in 0 0000 OK Cancel Setup Help Feature Size microns The nominal diameter of spots printed on the slide Sampling The level of detail you want to see in the image Select High for more detail Low for less detail and Optimal for normal detail Recommended Resolution The
106. on of all image pixels around the X axis e Flip X mirrors the position of all image pixels around the Y axis e Rotate rotates the position of all image pixels 90 counter clockwise 04 720046 000 Rev D 0804 40 arrayWoRx Standard amp Auto User s Manual Select Slides to Scan arrayWoRx Auto Scanner With the arrayWoRx Auto scanner slides are automatically selected for scanning after a cassette is loaded into the scanner A description of each slide in the cassette is displayed on the Scan Table The toggle buttons in the Scan column indicate whether slides are selected for scanning Note The arrayWoRx Auto Scan table displays the loaded slots at the top of the table For example if slots 2 4 and 6 are the only loaded slots in a cassette the table displays slots 2 4 and 6 followed by slots 1 3 5 and 7 In the following figure two of the occupied slots are selected for scanning Figure 13 Selecting Slides to Scan f FERES Oa Leland SHIQEU E Click this button to select de select a ae eat aa poas ls mr pan pan la pas a pia a To deselect slides e Deselect the Scan column toggle buttons on the Scan Table for the slides that you do not want to scan Deselected buttons change from blue to gray To deselect all of the slides e Click the Disable button to deselect all of the slides in the table TIP A quick way to select a few slides is to click the Disable button to deselect all of t
107. open image file 74 permissions 125 size required for scan 86 File Manager Gnome 112 File menu Image window 92 Main Scan window 74 File Names using bar codes 39 File Prefix changing 103 Filters calibration status and values 77 choosing a filter set 36 description 9 selecting 81 supported 144 Flat field calibration defined 63 refining 83 when to perform 63 Flattening stitch panel 39 Flipping TIFF image 39 Floppy disk drive mount and unmount 113 Fluor choosing filter set for 36 Fluorophore 8 Focus illustration 61 scanner 60 Focus Access door 60 Focus knob 61 Foot icon Gnome 110 Frequency of electric supply 11 Fuse 11 replacing 138 G Gain simulating laser PMT scanner 39 04 720046 000 Rev D 0804 Index 151 gedit 118 Get Single Panel 75 GIMP about 115 opening a file 116 Gnome 107 desktop 110 File Manager 112 LinuxConf 119 log on and off 18 108 main menu 19 110 mount drives 113 online Help 109 opening a TIFF 115 root user 119 screen capture 117 setting window options 124 task buttons 110 text editor 118 unmount drives 113 User s Guide 109 Gnome Documents 109 Gnome Panel starting scan from 19 Gnome User s Guide 109 Graphs 3D Intensity Profile 98 intensity histogram 100 Grayscale mode selecting 93 H Hard drive 10 Hazardous Voltage warning 14 Heavy Objects caution 14 Help Gnome 109 Image window 95 Main Scan window 77 online
108. or move a file 1 Find the file in File Manager 2 Right click the file that you want to copy to open a menu of file options jusrlocal softWoRx data sample_images ea gt File Edit Settings Layout Commands Help fat a amp erto R2 Rescan Home Icons Brief Detailed Custom Open with View My Scan_w595 tif Edit 7 oN 1o Sits MyScan Move to Trash ec sos sampleimagel ste calibration_files Delete tif G3 config aae Bi E data g i T sample_images symlink Ean ae eScanriles Scale20C j 000_w6 Scale200500_w59 ft ge Properties if 5 tif Cah 2 123 540 ee in 1 file Show all files 3 Choose COPY or MOVE from the menu Type or choose the name of the directory to which you want to move the file or Type the name of the file to which you want to copy the file 5 Click OK Tip You can also move a file by opening a File Manager window for the source and destination directory and dragging the file from the source to the destination directory To delete a file 1 Find the file in File Manager 2 Right click the file that you want to delete to open a menu of file options 3 Choose DELETE 4 Click Yes Working with the Floppy Disk and CD Drives Setting up a Linux system to use a floppy or CD is more involved than setting up these devices for a Windows system In Linux devices such as the floppy disk drive and the CD read write drive are represented as files Before Linux ca
109. ork settings in order to network the workstation Eth0O is the external network connection It is configured by default to use a static IP address of 172 20 20 20 with a related etc hosts table entry If you want to connect the Linux OS to your network you can use Linuxconf with root privileges to either reassign the static IP address to one that has been defined for your local network typically done by your sysadmin or to set EthO to DHCP for automatic IP address assignation The physical connection for Eth0O is labeled Network at the network cable interface port on the back of the computer Eth1 is the connection to the scanner It is configured to use a static IP address of 159 159 159 2 The physical connection for Eth1 is labeled Scanner at the network cable interface port on the back of the computer AN CAUTION The Eth1 setting should NOT be modified Modifying this setting causes communication to the scanner to fail Windows Configuration Windows uses Network and Private Virtual to communicate via your local network and to connect to the shared data directory in Linux Network is the external network connection It is configured by default to use DHCP to connect to your local network The physical connection for Network is labeled Network at the network cable interface port on the back of the computer If desired this ppliedPrecision Chapter 2 System Overview 13 network connection can be assigned a stat
110. out Mouse Shortcut You can also use the center and right mouse buttons to zoom in and out and center the image on the point you click Press the center button to zoom out the right button to zoom in Zoom to One Return the zoom level to the default zoom level ppliedPrecision Chapter 8 Image Window Reference 89 Status Bar The Status bar displays pixel intensity values and the zoom level When the mouse pointer is on the image the Status bar displays the intensity of each channel for the pixel that is under the mouse pointer When the mouse pointer is off the image the status bar displays a zoom level that indicates whether the image is larger or smaller than its original size An image displayed at its original size has a zoom level of one A zoom level greater than one indicates the image is zoomed in a level less than one indicates it is zoomed out Scale Bar Provides a scale for the image Scroll Bars Use the scroll bars to move the image horizontally or vertically in the Image window Click a scroll bar arrow to move the image incrementally or drag a slider to move more quickly The Image Window Buttons The Image window buttons allow you to navigate the Image window and adjust the intensity of the display lmel Scale Opens Scale Image that gives you control over the intensity scaling of the display in the Image window You can define the intensity envelope of the display for each channel and compare minim
111. page 34 Scan Starts a full scan of the scan area Equivalent menu command Scan NORMAL SCAN See Start and Stop the Scan on page 41 Stop Stops either a Preview scan or Normal scan that is in progress If more than one scan is remaining you can choose to cancel all remaining scans or only the current scan Equivalent menu command Scan CANCEL SCAN Eject Removes any slide loaded in the arrayWoRx Auto scanner returns it to the cassette and then ejects the cassette or removes any slide loaded in the arrayWoRx Standard scanner GppliedPrecision Bar code or Label Chapter 7 Main Scan Window Reference 79 Slide View Slide View in the Main Scan window is a graphical representation of the slide The region at the far left of Slide View represents the no scan area of the slide where a barcode or other label would normally be placed on the slide While arrayWoRx is scanning the file name of the current scan is displayed above the Slide View window When arrayWoRx is not scanning the number of the loaded slide is displayed Figure 28 Slide View Scan Area Rectangle The solid usually blue rectangle that you can use to define the scan area Panel Tool The square usually red that represents a panel A panel is the image acquired during a single exposure of the camera The image is made up of stitched together panels When the mouse pointer is over the Slide View The XY position of the mouse pointer is di
112. ply Layer Mask Apply Layer Mask Blend Blend Blinds The Blinds Filter Blur The Blur Filter The IIR Gaussian Blur Filter The RLE Gaussian Blur Filter Border Selection Border Selection Ck p yard Index introduction 4 116 arrayWoRx Standard amp Auto User s Manual Figure 45 An Image Displayed in a GIMP Window MyTIFF_685_595_ xtif 0 0 RGB 25 Ka Eafe A TP e012 1192 MyTIFF_685_595_xxtif 0 0 RGB 25 Cance To open a tif file in the GIMP e Double click the file in File Manager or e Choose PROGRAMS GRAPHICS THE GIMP from the Gnome Main menu choose File OPEN in the GIMP Toolbox and select the file from directories displayed in Load Image To close a file 1 Right click the GIMP image window to display the File menu J File d Edit _ New Select Open View save Image s Save As N Layers Revert Tools Mail Image Dialogs Print Filters Close Video Quit Script Fu E gt Q 2 Choose CLOSE To quit the GIMP e Choose File QUIT in the GIMP Toolbox Or e Right click the GIMP image window and choose Quit GppliedPrecision Chapter 10 Introducing Linux and Gnome 117 Capturing a Screen or Window in Gnome This section describes how to use the Screenshooter to capture an image of the screen or of a window on the screen There are several other ways to capture screens including the Linux import command For help with t
113. port if necessary UL 3101 1 First Edition Underwriters Laboratories Inc defines POLLUTION DEGREE 2 as Normally only non conductive pollution occurs Occasionally however a temporary conductivity caused by condensation must be expected 04 720046 000 Rev D 0804 12 arrayWoRx Standard amp Auto User s Manual Workstation Network Configuration This section describes the system s default network settings as delivered from the factory and how to modify these settings in order to connect the workstation to your network The workstation contains two network ports One is used for standard network communications and the other is used to communicate with the scanner Specific configuration settings are required to allow networked data transfer and scanner operation Note If you do not understand any of the terms in the following directions or you do not know how to obtain local network information please seek help from your IT Department before proceeding Linux Configuration Linux uses the EthO and Eth1 Ethernet connections to communicate via your local network and to operate the scanner We recommend that you connect to your local network via Windows running in VMWare see Windows Configuration below Files that you want to move to and from Linux can be saved in the shared_data directory and transferred or printed via standard Windows networking protocols Note It is not necessary to change the default Linux Eth0 netw
114. r finishing working in LinuxConf Additional information about using LinuxConf is available in the LinuxConf online Help system You can view parts of the help system by clicking the Help buttons in Linux Conf or you can access the entire help system GppliedPrecision Chapter 10 Introducing Linux and Gnome 121 Changing a User s Password Before changing a user s password you must log on to the system as root and open LinuxConf If you do not know the root password contact your system administrator and have them perform the following procedure for you To change a user s password 1 Ifthe Config tab in LinuxConf is not already on top click the Config tab 2 Inthe directory tree go to User accounts under Users accounts Normal This will display the Users accounts tab 3 Click the user name in the Accounts list on the Users accounts tab Click Passwd at the bottom of the User Information tab Config Tab Users accounts User information Pick a help file You must specify at least the login name and the full name User accounts Base info Params Privileges Group definitions Change root password M The account is enabled Special accounts Policies Login name awuser File systems Miscellaneous services boot made Full name PF ee group awuse i Supplementary groups Po Home directory opt Jhomefawuser 0 Command interpreter opt binasa O User ID opt 500 Quit Act Changes Help Accept Cancel Del
115. r status box before proceeding 8 Open mnt cdrom in File Manager to ensure that the files have been copied to the CD 9 Click Eject Disc and remove the CD from the drive If the CD does not eject right click on the CD icon on the Linux desktop and choose Unmount Volume from the popup menu 10 Choose File QUIT to close the Archive Manager 11 Close the File Manager windows Note If you copied files from a network to the workstation hard drive before copying to the CD you can use File Manager to delete the files from the hard drive after copying them to the CD 04 720046 000 Rev D 0804 106 arrayWoRx Standard amp Auto User s Manual ppliedPrecision CONTENTS Overview Logging On to and Off of Gnome Getting Help on Gnome The Gnome Desktop Working With Windows Managing Files with the Gnome File Manager Working with the Floppy Disk and CD Drives Using the GIMP to View TIFF Files Capturing a Screen or Window in Gnome Editing Files with the gedit Text Editor Managing User Accounts Using LinuxConf Changing a User s Password Deleting a User Account Creating a New User Account File Permissions Changing the Workstation Time Date Guidelines for using Linux 10 INTRODUCING LINUX AND GNOME This chapter is an introduction to the Linux operating system and Gnome user interface It describes the basic tasks you need to know to use the software The chapter also describes how to get more help from the Gnome online He
116. rag the blue scan area rectangle to the edge of the Slide view area To drag the rectangle position the mouse pointer inside the rectangle until it changes to Then move the mouse pointer 3 Find the boundaries of the scan by clicking on a point in the Slide view area that is outside of the scan area rectangle Clicking on a point moves the slide and acquires a preview image of that point This allows you to find the corners of the scan area 4 Position the blue rectangle so that it encloses the scan area To resize the rectangle point to one side with the mouse until the pointer changes to K Then move the mouse pointer If you are defining the scan area for a batch of slides leave some space between the blue rectangle and the array to account for variability in array printing Figure 10 The Slide View Slide View Slide 7 Drag a side of the scan area rectangle to enclose the array BE OLA BE Ate ALOR ees a ESTEN ETAS TATE E EE Jes Peres ame ip 4 ee OSS S25 5 Make sure that the desired channel is marked as active in the Channels tab To define a scan area from a full preview 1 Make sure that that the Status light is shining steady green indicating that there is a slide in the scanner Choose Entire Slide in the Preset list Click Preview and wait for the preview image to appear in the Slide View Aw N In the Slide View drag the sides of the blue scan area rectangle so that the rectangle enc
117. ragging its icon from a File Manager window into the gedit window To save and close a file in gedit 1 Choose File SAVE 2 Select the directory in which to save the file in the Save As window 3 Type the name of the file in the Selection field at the bottom of the Save As window Click OK 5 Choose File EXIT ppliedPrecision Chapter 10 Introducing Linux and Gnome 119 Managing User Accounts This section describes how to create and delete a user account and how to change a user s password using LinuxConf Be very careful of the changes that you make using LinuxConf You can make changes to the system configuration that will cause Linux to operate erratically or not at all Changing a user s password and deleting a user s account are straightforward tasks that you can accomplish with little experience with Linux and Gnome Adding a new user account requires that you make changes to the window manager settings that require experience with the Sawfish window manager Please contact the Applied Precision Hotline if you need assistance adding a new user account You can also manage user accounts from the Linux command line To display the help files for the command line commands open the terminal emulator and type man useradd man userdel orman passwd Note To change user account information you must know the root password to log on to Gnome as root 04 720046 000 Rev D 0804 120 arrayWoRx Standard amp Auto U
118. rated If the Calibration State displays Stale unload and reload the slide and try again If the Calibration State continues to display Stale call Applied Precision technical support Repeat Steps 7 9 for each filter set to be calibrated When the calibration is finished click Done to close Calibrate Scanner and eject the purple calibration slide The Scanner Lights 23 ppliedPrecision CONTENTS Overview Displaying an Image File in the Image Window Scaling Display Intensities Measuring Feature Dimensions Examining Intensity Data with Data Inspector Selecting a Region of Interest in Data Inspector 6 VIEWING IMAGE DATA This chapter describes how to do some common tasks using the Image window See Chapter 8 Image Window Reference for a complete description of the Image window features 04 720046 000 Rev D 0804 66 arrayWoRx Standard amp Auto User s Manual Overview The Image window is a set of tools that allows you to look at data stored in an image file You can look at a picture of the array a table of intensity values a three dimensional plot of intensity values and a histogram of intensity values You can display the information in these four formats simultaneously and easily move from one to the other to examine the data in the image file Figure 21 The Image Window 5 x ppliedPrecision Chapter 6 Viewing Image Data 67 Displaying an Image File in the Image Window
119. rce is a high intensity metal halide bulb and optical components that deliver a beam of white light to the excitation filter The light generated by the bulb is a mixture of visible and ultraviolet wavelengths The filter assembly holds up to four pairs of excitation and emission filters One emission filter and one excitation filter are required to measure the intensity of a single fluorophore Scanning software and motion control hardware place the appropriate filters in the light path for the fluorophore being measured A tube lens with a focusing mechanism focuses the image collected by the objective into the CCD camera The focusing mechanism is manually adjustable The CCD camera or Charge Coupled Device camera collects light from the emission filter and converts it into a digital signal that the scanner software processes to create an image file The CCD chip is an array of semiconductor devices or camera pixels Each camera pixel stores an electrical charge generated by light from the emission filter This charge is proportional to the intensity of the light or number of photons that reaches the pixel Electronic circuitry on the camera converts pixel electron counts from the CCD chip into a digital signal that represents the intensity of each pixel 04 720046 000 Rev D 0804 10 arrayWoRx Standard amp Auto User s Manual Workstation Overview The workstation is a personal computer that runs the Linux operating system It provides
120. re is a cassette in the cassette tray when the scanner enters Cassette Idle Mode the scanner pulls the cassette inside closes the door and inventories the slides If there is no cassette in the tray the scanner pulls the empty tray inside and closes the door To exit from Cassette Idle Mode press the Load Eject button on the scanner Note In Cassette Idle Mode the Status light shines amber if there is a cassette loaded in the scanner It remains off if there is no cassette loaded in the scanner Use Preview to Define the Scan Area A preview scan is a single channel low resolution scan that generates an image of the slide that you can use to locate the array s position Preview scans can be performed while the camera is cooling and the lamp is warming up This section describes two methods for using Preview to define a scan area that encloses the array e Define the area based on a partial preview when you know the approximate location of the array on the slide This method 1s faster than obtaining a full preview e Define the area based on a full preview when you do not know the location of the array or when the array is randomly positioned on the slide Scanning a Slide Multiple Times 43 ppliedPrecision Chapter 4 Scanning Slides 35 To define the scan area from a partial preview 1 Make sure that that the Status light is shining steady green indicating that there is a slide in the scanner 2 Click the Area tab and d
121. ring Intensity Cleaning Slides 11 MAINTAINING THE SYSTEM This chapter describes procedures required to keep the system hardware in optimum operating condition Electrostatic Discharge ESD There is a risk of damaging electronic components with electrostatic discharge when you are performing the maintenance tasks described in this chapter The following guidelines will help you avoid electrostatic discharge Electrostatic discharge is the release of static electricity from one place to another Static charges are easily generated and can be stored on people and various materials including plain plastics People often carry charges of 1000 to 5000 V but do not feel the sensation of a discharge under 3000 V Plastics can hold charges of several hundred to 25 000 V A few hundred volts can damage most semiconductor devices in a fraction of a microsecond e Keep static generating materials i e cardboard plastic sheets paperwork and tape three feet one meter away from the system e When you turn off the scanner keep the power cord plugged in so that the system 1s electrically grounded A CAUTION The system contains microelectronic devices which can be damaged by electrostatic discharge 04 720046 000 Rev D 0804 130 arrayWoRx Standard amp Auto User s Manual Cleaning the System Wipe the surfaces of the scanner as needed with a lint free cloth slightly moistened with water Replacing the Lamp Assembly The
122. s root this is required to load the files Insert the floppy containing the Service script in the workstation drive Right click on the floppy icon and choose Mount device this mounts the floppy drive Right click the floppy icon and choose Open Double click the install sh file this loads the files on the floppy disk Right click the floppy icon and choose Unmount Device Remove the floppy and log off of root ppliedPrecision Chapter 12 Troubleshooting 141 To send files to APLLC Service e After running the Awe Service Script choose one of the following options to send the service tar Z file inthe temp service directory to APLLC Service for review Send as an attachment to e mail Attach the the service tar Z file to an email and send it to servicehotline api com This is the recommended approach because it is the quickest method Also attach any image files that you want the technician to review If the image files are too big for you to email send them to the Applied Precision Web site as described below Send to the Applied Precision Web site Go to the http www appliedprecision com arrayworx serviceupload upload asp Web page Then follow the instructions on the form to send the service tar Z file and any image files that you want the technician to review Send in CD format Save your image files and the service tar Z file ona CD Then send the CD to Applied Precision LLC Inc 1040 12 Avenue N W Issaq
123. scanning using the currently specified Scan Settings preset The Scan Table displays the Slot Scan Preset and Data Name for each detected slide The tab also includes the Load Slide list Multi Scan Disable and Reset buttons See Also Scan Table 84 04 720046 000 Rev D 0804 32 arrayWoRx Standard amp Auto User s Manual Click here to list all slots available in Figure 9 Scan Table Tab cassette Click here to enable multiple scans of same slide a eae __ 4 __ 4 oat 4 ol ea pannia Slot The slots in the cassette containing detected slides Note The arrayWoRx Auto Scan table displays the loaded slots at the top of the table For example if slots 2 4 and 6 are the only loaded slots in a cassette the table displays slots 2 4 and 6 followed by slots 1 3 5 and 7 Scan Toggle button that determines whether or not the slide will be scanned Click the button to toggle the setting Blue indicates that the slide will be scanned gray indicates that the slide will not be scanned Preset Displays the default preset defined for the slide and allows different presets to be selected for each slide or multi scan Data Name Displays the slide identifier If the slide identifier is being read from the bar code on the slide the field cannot be edited If the slide identifier is not being read from the bar code this field will display a default slide name which can be edited that corresponds to the slot name A
124. senovedesnseusssensoueseonses pA Closing the Main Scan Window secimi En a a e E ZA Turning Off the Scanner and Workstation cccccccccccccccccccceececeeeeceeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeees 22 Thescainner Lieh eieaa a eds use es 23 4 Scanning Slides 25 Handling Slides With array WoRx Standard ccccccccccccccccsssccsevsccesesscecessseeesssseeesenseeeeenseeeeeaes 26 Insert the Slideinto the Scanner srini ianea iEn a EEA AE r a T a 26 Handling Slides With array WORK Auto ssseeeessssseesesssesssssssrressssreesssseressssressssereesssereessserressse 27 Insert the Slides into the Cassete sirana a a e aia enced ee Zi Load the Cassette INOS SCAMMER eraio e a a a Ea a e aia 30 N WS Wy IV SGC UAW sranani iane a a N N 31 Cassete VANS ModE eraat a a a e had sedeMeavemnast 34 Use Preview to Define the Sean ATE a ienen i a a a Ma ae 34 Choose he Filter and Bx POS URS eiri a e a baad 36 Choose the Resolution and Sensitivity airenisniianaieiid i a EE E A 37 Defne the Ady ance ODHONS senrenssiaena cutoemnandateeusraeiclld dest 38 Adyanced ab Op Ons eireiaeirai cai daar cuties aiid ies cea teamrenasid a cuntanitlateddeacuntanttrandd a aiuatuciaald dndse 38 Select Slides to Scan arrayWoRx Auto Scanner ccccccccccesssccceessssccceeesnsseeceeessseeeeeeessseeeeeens 40 boady Unload a Sim Ole Slide rennan aa aaan a ae 40 Sarrand SOPE SCA i a a tid ae lSo ierenseee aenonaeeS 41 Scannine a slide Multiple TIMES terdiiiernane akpi E A dads haeacoeeadh
125. ser s Manual Using LinuxConf LinuxConf is a Gnome user management tool that allows a system manager or user logged on as root to create and delete user accounts and change a user s password Note To use LinuxConf you must be logged on as root To open LinuxConf 1 Turn on the system 2 Type root in the Login box to log on to Gnome as root If you do not have access to the root password contact the Applied Precision hotline see page 6 3 Type the root password in the Password box Choose PROGRAMS SYSTEM LINUXCONF from the Gnome Main menu to open LinuxConf Figure 46 LinuxConf awe_tech Linuxconf1 19 subrey 2 Config Control Status Users accounts User information Pick a help file You must specify at least the login name Networkin Networking and the full name E Users accounts Normal Directory Tree 4 Group definitions Base info Params Privileges Change root password M The account is enabled Special accounts Policies Login name awuser File systems Miscellaneous services boot mode Full name ee group awuse i Supplementary groups PO Home directory opt Jhomefawuser 0 Command interpreter opt binasa O User ID opt 900 Quit Act Changes Help Accept Cancel Del Passwd Tasks Help A CAUTION When you are logged on as root you can damage the system configuration Do not make any changes that may have effects that you are not certain of Always log off afte
126. sing windows on the Gnome desktop The behavior of your desktop depends on the Sawfish window manager settings Making a window active causes it to be on top and makes the buttons menus and fields in the window respond to the mouse To make a window active e Using the middle mouse button click anywhere in the window When you minimize a window the window disappears from the desktop but the application continues to run To minimize a window e Inthe top right corner of the window click the minimize box elias ex To open a minimized window 1 Locate the task button for the window task bar at the bottom of the desktop 2 Click the window s task button Terminal Ee Netscape Welcomgy 0 arrayWoRx e Scan aor The GIMP Task Buttons Note The best way to close a window is to close it from the File menu Using the close box to close a window may not stop all processes associated with the application running in the window To close a window e From the File menu choose CLOSE or QUIT Sometimes an application stops working properly and does not respond to the mouse To close an application that has stopped responding 1 Right click the application s button in the task bar 2 Choose KILL APP 04 720046 000 Rev D 0804 112 arrayWoRx Standard amp Auto User s Manual Managing Files with the Gnome File Manager This section describes how to open copy move and delete files using the
127. sion filter to ensure that the camera sees only a narrow band of wavelengths that includes the emission wavelength of the fluorophore under investigation The CCD chip is an array of pixels When the chip is exposed to light 1t generates a 14 bit value for each pixel This value represents the number of photons that reach the pixel during the exposure Camera electronics and scanner software convert this photon count into a 16 bit value that is the intensity value for the pixel on the image Signal to Noise Ratio Because signal variation noise can significantly reduce data quality you ll need to set the exposure resolution and sensitivity to maximize the Spot SNR signal to noise ratio This ratio should not be confused with the signal to background ratio commonly referred to in microarray experiments For more about Spot SNR see Appendix B Signal to Noise Binning Binning is a technique used to combine information from adjacent pixels on the camera into one pixel on the image Binning increases the number of photons used to generate a single image pixel and thus increases the signal to noise ratio In 2 by 2 binning for example the intensity values of four camera pixels are used to calculate the intensity of a single image pixel Binning can either occur on the camera hardware binning or in the scanner software software binning The Exposure Resolution and Sensitivity settings on the Main Scan window give you control o
128. sity of the display The Scale Pan Center and Home button descriptions begin on page 89 Wavelength Selector Click one of the two wavelengths in the Wavelength Selector to show or hide the wavelength in the Image window You can show one wavelength at a time or both simultaneously For example the 595 nm channel can be shown in green the 685 nm channel in red or both simultaneously See View Menu on page 93 for a description of how to change display colors Toggle the display of a channel by clicking the wavelength in the Wavelength Selector Click once to select the wavelength yellow box Then click again to turn the channel off or on Intensity Quick Scale Drag the Quick Scale slider up or down to increase or decrease the maximum intensity displayed in the Image window for the channel highlighted on the Channel Selector See Scale Image on page 90 for a more detailed description of display intensity Zoom Wheel Drag the Zoom wheel up to zoom out make the image smaller or down to zoom in make the image larger Note that you can move the pointer anywhere on the screen while zooming in or out You can incrementally zoom in and out by clicking the zoom arrows directly above and below the zoom wheel Click the up arrow to zoom out the down arrow to zoom in TIP You can also control the zoom with the mouse Click the right mouse button on a point to zoom in and center the image on that point Click the middle mouse button to zoom
129. sity values at or above the maximum are displayed as the same value e g 255 or white Click Scale in the Image window to open Scale Image Note Changing intensity scaling values does not alter image data only the display Figure 37 Scale Image Scale Image Window 1 Wave 595 v 685 Display Min Max Exp 35 88 2302 91 1 00 Image Min Max 35 88 11979 8 q JT Show Histogram 7 Log Number of pixels y axis Intensity x axis y Uone Restore Default Scale Hely Minimum Image Maximum Intensity Image Intensity Min Max Display Intensity Handles You can drag a Min Max display intensity handle left or right to change the display intensity The Max Display Intensity handle defines the upper cutoff intensity the display intensity of the brightest pixel The Min Display Intensity handle defines the lower cutoff intensity the display intensity of the dimmest pixel In general decrease the upper cutoff intensity to make spots appear brighter and increase the lower cutoff frequency to make spots appear dimmer Note You should scale an image before you save it as a 24 bit RGB image Wave The channel to scale is linked to the Channel Selector in the Image window You can choose a channel by either clicking a Wave value in Scale Image or by selecting a channel in Channel Selector Scale Image displays information about the channel indicated by Wave For example the intensity values in Figure 37 are for the
130. software calculates and displays a resolution value based on the Feature Size and Sampling that you entered To use the Resolution Calculator 1 Open the Resolution Calculator 2 Type a value in Feature Size which is the typical diameter of a circular spot on the slide 3 Select a Sampling level 4 Click OK to automatically enter the suggested value in Resolution on the Image tab 5 Click Done to close the Resolution Calculator To customize the Resolution Calculator 1 Open the Resolution Calculator and click Setup 2 Enter values for each Sampling level Each value is the number of pixels that will be used to represent a single spot or other feature in the image 3 Click Done to close Resolution Calculator Setup See Also Resolution 53 Measuring Feature Dimensions 69 GppliedPrecision Chapter 5 Refining Scan Settings 57 Slide and Microarray Dimensions The scanner accepts standard microscope slides The array must be printed within the area enclosed by the dotted line in Figure 18 to avoid interference from the slide edge during a scan and to provide a surface for the slide holder to grip the slide The allowable area for the array to be printed on the slide also takes into consideration the space needed on the slide for a bar code or other label A bar code or other label should be placed on the side of the slide on which the array is printed CAUTION Slides that do not meet the following requirements
131. splayed in the status bar The lower left corner of the Slide View is the origin Note The values in the text boxes on the Scan Area tab change as you move or resize the scan area rectangle Another way to set the size and position of the scan area is to type values in the text boxes 04 720046 000 Rev D 0804 80 arrayWoRx Standard amp Auto User s Manual Scan Settings Preset A preset is a set of scan settings that you can define for use with slides that require the same scan settings A preset is selected for each slide in the Scan Table The preset scan settings include e Scan Area e Number of Channels e Exposure e Resolution e Sensitivity e Generate TIFF Images e Gain Settings Figure 29 Scan Settings Preset Scan Settings Preset Sample E Save Create Delete Panel View Scan Table Preset The name of the set of scan settings Save Save the current settings as a set with the name shown in Preset Create Save the current settings to a new name Delete Delete the set of scan settings whose name is shown in Preset Important You cannot delete a preset if it is being used by any of the entries in the Scan Table The software will display a warning message and prevent the deletion See Also Working with Presets 48 GppliedPrecision Chapter 7 Main Scan Window Reference 81 Scan Settings Tabs The Scan Settings Tabs together with Slide View are where you choose scan settings before s
132. t if the drive is mounted Tip The CD will mount automatically when it is first inserted To unmount a drive 1 Choose PROGRAMS SYSTEM DISK MANAGEMENT on the Gnome Main menu 2 Click Unmount for the drive you are mounting in the User Mount Tool e mnt floppy to unmount a floppy disk e mnt cdrom to unmount a CD Tip If there is an icon on the desktop for the drive you can mount and unmount it by clicking the icon with the right mouse button and choosing either Mount or Unmount from the menu GppliedPrecision Chapter 10 Introducing Linux and Gnome 115 Using the GIMP to View TIFF Files The GIMP is an image manipulation program When you open a tif file from the File Manager it opens in the GIMP You can open the GIMP by choosing PROGRAMS GRAPHICS THE GIMP from the Gnome Main menu The GIMP displays the image in one window and the Toolbox and other tools in separate windows Figure 44 The GIMP Toolbox To get help from the GIMP online Help system 1 Choose Help HELP in the GIMP Toolbox 2 Click the Index tab in the GIMP Help Browser Index 04 720046 000 Rev D 0804 eats GIMP Help Browser lt Back gt For Index About About Dialog Add Alpha Channel Add Alpha Channel Add Layer Mask Add Layer Mask Airbrush Airbrush Alpha Channel Add Alpha Channel Alpha to Selection Alpha To Selection Alpha to Selection Anchor Layer Anchor Layer Apply Canvas The Apply Canvas Filter Ap
133. t date 43 Remove Cassette from the arrayWOoRx Auto Scanner ccccccccceessccceeessseecceeessseeceeesseeeceeeesaes 44 Vew DEFIE npare r a T OE NOE CREE Co eee 44 04 720046 000 Rev D 0804 IV arrayWoRx Standard amp Auto User s Manual Refining Scan Settings Choosing Scan OPtions cccccceceeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeeees Working witht Presets ennaa ai Exposure Resolution and Sensitivity ccccccccccccccccceeeeeeeeeeees ER DOSU Oaa a a E ahaha E RESON seee a a E EA 6 1 T aae a ne en NEA Displaying Pixel Intensity in Panel View ce eeeeeeeeees Using the Resolution Calculator cc ccccccccceeeeeeeeeeeeeeees Slide and Microarray DimensiOD3 0cceeeeeeeeeeeeeeseeeeeeeees Extending the Scan AAT Chick Secs oh hs tak a a ahaa accra Using Deconvolution sca hc esate E ete cated POCUSING th Scanner sitet forte hota a a cated Flat Field C aM bat oMescsci wens tess ode deceidte ted aaeaseedsnsauaet Viewing Image Data ON TVO Wy aca becathcs het dew Nace Seecncdta a esedaecdeeeip Meaiesasasiadl Displaying an Image File in the Image Window Scaling Display Intensiti Ss sa tas tse sesshgndacteas sahara dla cumnsueg ceaets Measuring Feature Dimensions cccccccccccceeeecceeeeeeeeeeeeeees Examining Intensity Data with Data Inspector Selecting a Region of Interest in Data Inspector Main Scan Window Reference Fle MeO sas se aes so execs wate a A EEA nee T Sean MEN eE
134. tem is equipped with safety labels and interlocks to help protect you from potential injury that may be caused by ultraviolet light heat or the high voltage of the scanner s lamp Ultraviolet Light and Heat The following hazards are associated with the scanner lamp assembly s high intensity metal halide bulb e Damage to eyes and skin caused by exposure to ultraviolet radiation e Burns caused by contact with a hot lamp assembly e Fire ignited by hot lamp assembly e Interaction of nearby chemicals with ultraviolet radiation e Damage caused by placing apparatus too close to the scanner To avoid injury or damage do not remove the scanner cover Also keep chemicals and equipment that are sensitive to ultraviolet radiation away from the scanner Note The lamp access door and filter assembly are equipped with safety interlocks that turn off the lamp if the door is opened or the filter assembly is removed while the lamp is on WARNING Under certain conditions the bulb can explode If this occurs the fumes can be toxic Evacuate the room immediately and remain out of the room for at least 30 minutes Call Applied Precision for a replacement bulb assembly Class 1 LED Product Information A WARNING The arrayWoRx system is classified as a Class 1 LED Product IEC 60825 1 because it uses a bar code scanner with a red 660nm LED If the bar code scanner requires maintenance please contact Applied Precision ppli
135. th Zones Bar Chart F Legend Keep Scale Constant Add Optional Title M M M 119 1566 897 491 Total 297967 SD 352 44375 Print Help Close Rotate and resize the graph using the mouse and key combinations shown in Table 7 Table 7 3D Graph Rotation and Zoom Commands Action Mouse Key Rotate Use the middle mouse button to rotate the graph in any direction or Hold the X Y Z or E key and use the middle mouse button to rotate the graph around a single axis Move Hold the SHIFT key and use the middle mouse button to move the graph to a new position in the window Zoom Hold the CTRL key and use the middle mouse button to drag the graph up or down or to zoom out or in Zoom Region Hold the CTRL key and use the left mouse button to select a region of the graph and to zoom in on the region Reset Zoom Press the r key to return the graph to its original size Mesh Allows you to show or hide the mesh outline Shaded Allows you to show or hide the polygon surface Contours Allows you to show or hide iso value lines GppliedPrecision Chapter 8 Image Window Reference 99 Zones Turns on or off color coding of value ranges Smooth Zones Turns color blending on or off when Zones is selected Note To view a two dimensional contour map of the region of interest turn off Mesh Shaded and Barchart and turn on Contours Zones and Smooth Zones Bar Chart Allows you to toggle between bars and sur
136. the pixel in the Image window Line Profile Defines and plots an intensity profile graph of a row or column of pixel intensities in the Image window For example you can view a graph of pixel intensities for a row of spots You can set options to draw lines horizontally or vertically You can also move lines by clicking the left mouse button Data Inspector Opens the Data Inspector to view a table of intensity values and look at intensity data and basic ratio data in different ways See Examining Intensity Data with Data Inspector on page 70 Measure Distances Measures the size of features in the Image window such as spot diameter in nanometers microns millimeters or pixels See Measuring Feature Dimensions on page 69 Help Menu The Help menu command ON WINDOW opens the online Help topic for the Image window You can also open online Help topics by clicking Help in a dialog to display help about the options in the dialog 04 720046 000 Rev D 0804 96 arrayWoRx Standard amp Auto User s Manual Data Inspector Data Table The Data Inspector displays image intensity and intensity ratio data from a region of interest ROT in the Image window in various formats Using Data Inspector you can define the size of the region of interest and save data to ASCII files Figure 38 Data Inspector Data Table The data table is the table of values displayed in Data Inspector It displays intensity values of each pixel in t
137. tion software fails to launch Scanner is not functional Blown fuse The Give root password for maintenance prompt appears while Linux is starting up Buffer Underrun Cables loose disconnected or connected to Ethernet port on workstation instead of scanner port Improper shut down of acquisition software This may occur when Linux or Windows have not been shut down properly The CD recorder asks for its next serving of data and the data isn t ready This will most likely result in an unusable disc Check the Ethernet cables re ret 1 From the Linux Main menu choose PROGRAMS ARRAY WORX STOP ARRAY WORX 2 Wait a few seconds for files to be cleaned up Then try to re launch the acquisition software 1 Perform a visual inspection e g check for cables that are not connected 2 Shut down and then restart the workstation and scanner 3 If the system still does not work call the Applied Precision Service Hotline Use the 10A Slow Blow fuse that is shipped with the system for use with 100 120V If primary power is 220 240V use a T6 3AH fuse as an alternate Open a Linux Terminal window and enter the following commands gt umount mnt cdrom gt mount mnt cdrom 1 Type the root password and press ENTER 2 At the system prompt type fsck 3 At each Repair filesystem prompt press ENTER until you see the message System has been modified 4 At the system prompt type fsck
138. to make button labels and button tool tips visible or invisible and to enable the options on the Advanced tab See Buttons on page 78 Quit Closes the Main Scan Window See Closing the Main Scan Window on page 21 GppliedPrecision Chapter 7 Main Scan Window Reference 75 Scan Menu The commands on the Scan menu allow you to start and stop a scan and to choose options for the scan de Cancel Scan Ctrl C Get Single Panel Ctrl enyana Clear Preview Scan Options Preview Scan Starts a low resolution scan of the area defined in Slide View See Use Preview to Define the Scan Area on page 34 Normal Scan Starts a full scan Note If you are using arrayWoRx Auto and a slide is loaded when you start the scan you are prompted to choose whether to scan only that slide or to scan all of the slides that are selected in the Scan Table See Start and Stop the Scan on page 41 Cancel Scan Stops either a Preview scan or Normal scan that is in progress Note If more than one scan is remaining you can choose to cancel all remaining scans or only the current scan When you stop a scan that is in progress no data is saved from that scan However data collected from all previous scans 1s saved Get Single Panel Acquires the image of a single panel at the location on the slide indicated by the panel tool in Slide View See Slide View on page 79 Clear Preview Clears all images displayed in Slide View
139. to see 1f it is described in Chapter 4 Scanning a Slide or Chapter 5 Refining Scan Settings You can also find detailed descriptions of each feature in online Help Figure 26 The Main Scan Window Menus page 74 Buttons page 78 Slide View page 79 Panel View page 83 Status Bar page 86 04 720046 000 Rev D 0804 74 arrayWoRx Standard amp Auto User s Manual File Menu The File menu commands allow you to open files associated with the image file that is created when you scan a slide and to change the way buttons on the Main Scan window are labeled array WoRk e Scan File Scan Utilities Help Open Image File Ctr O show Data Folder Ctrl D Archive Data To CD Preferences 956909 90 49 Quit Open Image File Allows you to select an image file with the file extension stitch or stc and then open an Image window displaying the file you selected See Displaying an Image File in the Image Window on page 67 Show Data Folder Opens a Gnome File Manager window so that you can open files of any type Use this command to open TIFF files See Using the GIMP to View TIFF Files on page 115 Archive Data to CD Starts the ArrayWoRx CD Archive Manager which allows you to make copies of data files ona CD See Saving Files to CD on page 104 Important The Archive Data to CD function only works in Linux mode not Windows VMWare Preferences Allows you
140. tory Double click the diagnostic term sh file If your system doesn t have this file contact APLLC to get installation media and follow the instructions in To install the Awe Service script on page 140 Follow the directions in the terminal window At the Select Branch prompt choose the option specified by the APLLC technician e For Scan Client output verify that the main arrayWoRx toolbar is open Then enter c e For Main Toolbar output enter m Run the script as directed by the technician To stop verbose output close the arrayWoRx software application Follow the directions in the terminal window to gather the rest of the files If you are prompted to collect root access enter n unless the technician specifically requests these files If you choose to collect root access files you will need to enter the root password The Scanner Service files are collected and placed in the service tar z file in the tmp service directory This file is compressed and ready to send to APLLC Follow the instructions to install the Awe Service scripts if your workstation does not have the service term sh file or the diagnostic term sh file Before you start contact APLLC to get the files that you need to install these scripts To install the Awe Service script l ARE eet SS Load the five Awe Service script attached files onto a floppy drive Turn on the ArrayWoRx Workstation and Scanner Log on to the Workstation a
141. ts are still saturated change the sensitivity setting to Dynamic Range 9 Repeat steps 3 and 4 to verify that the spot is no longer saturated 10 Repeat steps 2 9 for the CY5 channel See Also Using the Resolution Calculator 55 Measuring Feature Dimensions 69 ppliedPrecision Chapter 5 Refining Scan Settings 55 Displaying Pixel Intensity in Panel View During a scan the colored bar at the bottom of the Panel view displays the channel and its minimum maximum and mean intensity values It also displays the intensity and percent of optimum values of the pixel that is under the mouse pointer The percent of optimum value is the intensity of a spot relative to the maximum intensity that the CCD camera can measure You can use these intensity and percent of optimum values to choose an exposure setting for the scan and avoid saturated spots in the image Figure 17 Percent of Optimum in Panel View Panel View Scan Table ES The arrayWoRx Percent of Optimum intensity calculation is based on the full dynamic range of the detector CCD camera By convention the optimum intensity is defined to be 80 of the full dynamic range At saturation the maximum limit of the dynamic range the calculated percent is 125 Image intensities above the optimum indicate at least one of the pixels in the detector is nearing saturation 125 of Optimum To avoid saturating areas of interest you may need to reduce the exposure time Image i
142. uah WA 98027 8929 Note If you have a problem running these procedures or need additional assistance please call the Customer Service Hotline at 1 800 862 5166 04 720046 000 Rev D 0804 142 arrayWoRx Standard amp Auto User s Manual arrayWoRx Problem Report Form Research Facility System Serial Number Contact Person softWoRx version use Help Software Version to display Problem Encountered Please write a detailed description answering as many of the following questions as possible Questions 6 Is the failure the same each time or does it show different symptoms o o o Z o o o 8 Does it go away after the workstation is re booted oS 9 Does it go away after the instrument controller is re booted o Z o o SSS Additional Comments See Page 2 for additional clarification issues Please e mail this form to hotline api com or Fax to 1 425 557 1055 attn Bio Service Hotline ppliedPrecision CONTENTS Filters and Dyes supported by arrayWoRx APPENDIX A FILTERS AND DYES array WoRx supports the filters and dyes shown on the following table If you would like to add third party filters to your arrayWoRx scanner contact Applied Precision Customer Service at 1 800 862 5166 or servicehotline api com 04 720046 000 Rev D 0804 144 arrayWoRx Standard amp Auto User s Manual Filters and Dyes supported by arrayWoRx Alexa 532 Alexa 546
143. ughly as possible before slides are scanned Use one of the following recommended methods to clean slides e Use the edge of another slide to scrape the back non array side of your slide e Using an anti static gun slowly squeeze and release the trigger several times approximately 10 cm away from the non array side of the slide e Use acombination of the above scrape the slide use the anti static gun and then scrape the slide again 04 720046 000 Rev D 0804 136 arrayWoRx Standard amp Auto User s Manual ppliedPrecision CONTENTS Problems with Images Other Problems Running the Service Script arrayWoRx Problem Report Form 12 TROUBLESHOOTING Problems with Images Condition Probable Cause Action Dim spots on Exposure time not long Increase exposure time image enough Seeing Rescan the slide with Apply Panel Flattening changed to the paneling opposite setting For example if Apply Panel Flattening is turned on and you see paneling try turning it off and rescanning the slide If paneling persists contact the Applied Precision Service Hotline for assistance Bulb may not be bright Check brightness measurement l If necessary replace bulb Bright spots on Dust on slide Clean slide image 04 720046 000 Rev D 0804 138 arrayWoRx Standard amp Auto User s Manual Other Problems Condition Workstation can t connect to the scanner acquisition window opens partially Acquisi
144. um maximum and mean intensity values of the display with the intensity values of the collected data stored in the image file The adjustments you make in Scale Image only affect the way the image appears on the screen they do not affect the data stored in the Image file See Scale Image on page 90 ay Pan Turns the pointer into a hand shaped pointer that you can use to drag a point in the image to a new position rod Center Turns the pointer into a circle that you can click on a spot in the image to place that spot in the center of the Image window et Home Moves the image to its position in the Image window before the last position change made using the Pan or Center button 04 720046 000 Rev D 0804 90 arrayWoRx Standard amp Auto User s Manual Scale Image r k Pal a 1 a Max Display Intensity Handle Min Display Intensity Handle The options in Scale Image determine how the intensity value of each pixel stored in the image file is mapped to a display intensity value in the Image window A scaling function determines the mapped values Scale Image gives you control over three parameters of the scaling function the minimum display intensity value the maximum value and an exponent value that affects how values between the minimum and maximum display intensity values are determined Intensity values at or below the display minimum are all displayed with the same display value e g 0 or black Inten
145. und intensity is a complex combination of chemistry slide autofluorescence and scanner imaging performance The baseline value of zero is essentially arbitrary among different scanners Comparisons of low intensity data should therefore be made using relative information such as measurements of Spot SNR values taken from the same spot Dynamic Range Yet another common misconception involves dynamic range Scanner B does not have twice the dynamic range of Scanner A simply because its background is 100 counts as compared to 200 counts To be meaningful dynamic range should be calculated using the lowest detectable spot as the value in the denominator not the average background intensity 04 720046 000 Rev D 0804 148 arrayWoRx Standard amp Auto User s Manual Conclusion Spot SNR provides a much more accurate measure of scanner performance than the signal to background ratio commonly and erroneously called SNR When a slide s chemistry is of high quality 1 e spot and background morphologies are very uniform duplicate measurements of Spot SNR can be made with different scanners to accurately compare imaging performance Without accounting for variation signal to background measurements should be avoided because conclusions made about image quality and scanner performance are usually erroneous and misleading ppliedPrecision Index 149 INDEX A Accounts user 119 Acquire Single Panel Image 84 Adjusting
146. ure Distances L pror SS Distance between start point and end point To measure the diameter of a spot t 1 Open an image file in the Image window Help 2 Zoom in on the spot that you want to measure 3 Choose Tools MEASURE DISTANCES 4 Click a point on the circumference of the spot to define the start point 5 Click a point on the spot s circumference directly opposite the first point to define the spot s diameter as shown in the previous figure This is the end point The length of the line you have defined is displayed in Measure Distances 7 To view the length in different units click the Units button and select the units that you want to use Note If you click the Units button after making a measurement you must re do the point selection see Also Displaying an Image File in the Image Window 67 04 720046 000 Rev D 0804 70 arrayWoRx Standard amp Auto User s Manual Examining Intensity Data with Data Inspector The Data Inspector includes several tools for examining the intensity data in the image file This section describes how to simultaneously view a graphical image table of intensity values 3 D graph and histogram of intensity values and move between these different formats Figure 24 Image Window with Data Inspector Tools To open the Data Inspector tools Open an image file in the Image window From the Tools menu choose DATA INSPECTOR In Data Inspector
147. ut saving the changes or click Cancel to return to the Main Scan window You have made changes to the current Scan Settings Do you want to save them 4 Inthe confirmation box click Yes to close the Main Scan window 04 720046 000 Rev D 0804 22 arrayWoRx Standard amp Auto User s Manual Turning Off the Scanner and Workstation CAUTION Before turning off the scanner and workstation you must close the Main Scan window as described in the previous section You must shut down the Windows operating system before you shut down or log off of the Linux operating system Do not shut down the Windows operating system by clicking the Close box in the top right corner of the VMWare Workstation window This can corrupt Windows system files and require non warranty service repairs To turn off the scanner and workstation 1 On the Analysis toolbar choose File Close to close the toolbar Then close any other windows and applications that are running on Gnome or Linux 2 Ifyou are running Windows close any windows or applications that are running on Windows Then from the Windows Start button choose SHUT DOWN In the Shut Down Windows dialog box select Shut Down and click OK When the VMWare dialog box appears press CTRL ALT and click Power Off Important You will not be able to regain use of the cursor unless you press CTRL ALT 3 Close any windows and applications that are running on Gnome or Linux 4 Clic
148. ve Data to CD function only works in native Linux mode not Windows VMWare 2 If the files you want to copy to CD are on a network drive we recommend that you copy the files to the hard drive on the workstation before copying them to a CD This will minimize potential data transfer delays that may make it difficult to create the CD To copy files to a CD using the CD Archive Manager 1 Place a blank CD R in the CD drive 2 Choose ARCHIVE DATA TO CD on the Main Scan window File menu or click the Archive Data to CD button on the toolbar to open the CD Archive Manager File Options Catalog Drag Files or Directories folders to the list area below Create CD Cancel Eject Disc Delete Selected Delete All CD File Name lt Source File Name 478 Mb 75 Full MyScan stc lt usr local sof tWoRx data sample_images MyScan stc MyScan_w595 tif lt usr local sof thoRx data sample_images MyScan_w595 tif MyScan_w685 tif lt f usr local sof thoRx data sample_images MyScan_w685 tif 3 Click Files to display the File Manager fustlocal softwoRx data sample_images Sj mix Eie Edit Settings Layout Commands Help da A b aa amp Bt gg Back Up Forward Rescan Home Icons Brief Detailed Custom Location usr local softWoRx data sample_images aa A My Scan ste MyScan_w595 tif DOTARE ie anA iL 3 sbin 2 a E Sy sotwoRx D Ga bin G calibration_files MyTIFF_685_595 Samplelmaget ste Sca
149. ver how the intensity value is determined from the photon count The settings determine e the maximum dynamic range of the image e the amount of memory required to store the image e the time required to scan the slide Choose settings that achieve the balance that your experiment requires between these factors Refer to the following sections for information about the effect of these settings on the scan 04 720046 000 Rev D 0804 52 See Also arrayWoRx Standard amp Auto User s Manual Exposure The exposure is the amount of time in seconds that a pixel of the CCD is exposed to the emission light from the excited sample The exposure should be set to e minimize saturation of bright spots e maximize signal to noise ratio e minimize the amount of time it will take to scan the slide In general the exposure time should be set to avoid saturation yet maximize the signal to noise ratio If the exposure is too long the images of the brightest spots will saturate and details on the spots will not be visible If the exposure is too short information about the dimmest spots will be lost Both the signal to noise ratio and scan time increase as the exposure time increases It is helpful to observe the percent of optimum in the Image view A value of 125 percent of optimum indicates pixel saturation See Displaying Pixel Intensity in Panel View on page 55 When determining the exposure setting be careful not to use image pa
150. w or hide 88 Web site Applied Precision 5 6 White light source 9 Window close 111 make active 111 minimize 111 Window manager Sawfish 111 Window Number 88 Windows DHCP 12 network configuration 12 switch to 18 VMWare 10 Workstation configuration 10 connecting to scanner 13 Z Zoom 3D Data Graph 98 Image window 88 93 Panel View 83 ppliedPrecision
151. ware This dialog contains three tabs General Filters and Advanced e General displays information regarding the system s serial number scan server version instrument controller version hardware controller OS scanner memory uptime and scanner disk usage e Filters accesses dialogs for viewing the wavelength bandpass calibration status and calibration values for each filter installed in the system e Advanced displays current camera temperature and bulb age Also accesses dialogs that allow bulb age to be reset log files to be downloaded from the scanner filter descriptions to be edited and bar code symbologies to be enabled Software Versions Opens a dialog that displays information about the software versions 04 720046 000 Rev D 0804 78 arrayWoRx Standard amp Auto User s Manual Buttons il _ The Main Scan Window buttons are shown in Figure 27 You can hide or show text labels on buttons using the PREFERENCES command on the File menu Figure 27 The Main Scan Window Buttons File Scan Utilities Help Data Opens a Gnome File Manager window so that you can open files of any type Equivalent to the File menu command SHOW DATA FOLDER See Managing Files with the Gnome File Manager on page 112 Preview Starts a low resolution scan of the scan area indicated by the Scan Area rectangle in Slide View Equivalent menu command Scan PREVIEW SCAN See Use Preview to Define the Scan Area on
152. wo primary sources the slide s chemistry printing and hybridization and the microarray scanner itself High quality printing and hybridization require high quality images to extract the maximum amount of useful data from the slide In other words low chemistry noise from careful optimization of spot printing and hybridization will be of limited benefit if scanner noise is high as shown in Figure 49 Noise also reduces the dynamic range because the spot detection limit is higher for noisier images the lowest detectable spots must be brighter to overcome the effects of noise reducing the range of useful intensities for the experiment Noise amp Spot Detection For microarray experiments a measure of spot vs background that also includes the effects of image noise can be derived as follows Spot SNR Mean spot Mean rackground Standard Deviation background This calculation can be called the Spot SNR Using this formula Scanner A in Figure 49 provides a Spot SNR value about 6 times higher than Scanner B due to much less noise in the image Adopting a standard detection criterion of Spot SNR 3 0 Scanner A readily detects the spot whereas Scanner B fails to detect it Background Figure 49 also illustrates another common fallacy namely that lower background intensities yield higher quality data In this case the lower average background intensity in Scanner B s image did not make the spot more detectable Backgro
153. x Standard and arrayWoRx Auto Scanners Scanner Overview The scanner components are the optical system the precision motion control system and the computer that is embedded in the scanner The optical system is based on a CCD camera and white light source The excitation filter is a band pass filter that selects a band of wavelengths from the full spectrum available from the white light source These excitation wavelengths are delivered to the sample on the slide by fiber optics The emission filter and other optics ensure that only the wavelengths emitted by the excited fluorophore on the sample reach the CCD camera Note Virtually all wavelengths in the visible spectrum can be scanned by the arrayWoRx scanners The precision motion control system positions each section of the slide precisely in the optical path to ensure that the images of the sections are properly aligned in the acquired image The embedded computer hosts the scanner software that controls the scanning process This computer is connected to the arrayWoRx workstation through a dedicated ethernet cable It can be controlled and configured from the arrayWoRx Main Scan window ppliedPrecision Chapter 2 System Overview 9 Figure 2 Components of the CCD camera Based Optical System CCD camera F White light source S Excitation filter A fluorescence _ Emission filter Fiber optic ring Slide The white light sou
154. x Mean intensities Status bar indicates scan progress To stop a scan e Press the Stop button in the Main Scan window Stop Scan Important Notes 1 When you stop a scan that is in progress no data is saved from that scan However data collected from all previous scans is saved 2 You cannot stop a scan while slides or data files are being transferred 3 If you are using the arrayWoRx Auto scanner the currently loaded slide will not be unloaded when you press the Stop button 4 If you are using the arrayWoRx Auto scanner to scan only one slide that slide will not automatically eject when the scan is complete ppliedPrecision Chapter 4 Scanning Slides 43 Scanning a Slide Multiple Times You can use the Multi Scan option to scan one slide several times This option can be useful in several different ways e You can scan only the areas of the slide that you are interested in and exclude areas that do not contain data This is accomplished by defining which areas of the slide to scan and performing a separate scan for each area e You can scan the same area several times with different settings e g exposure times pixel sizes resolution etc for each scan e You can define areas of a slide and scan each area with different settings Clicking the Multi Scan button on the Scan Table tab of the Main Scan dialog allows you to select a slide If only one slide is present it is automatically added to the
155. xperienced users who are looking for command syntax and parameter lists The Info pages are similar to the Man pages but provide more in depth information about the commands The Info pages are written for experienced Unix or Linux users Gnome Documents is a comprehensive list of the online Gnome documentation It is organized alphabetically by feature name When you can t find the information you are looking for in the Gnome User s Guide try looking in Gnome Documents 04 720046 000 Rev D 0804 110 arrayWoRx Standard amp Auto User s Manual The Gnome Desktop The Gnome Desktop is the workspace on the screen Figure 42 The Gnome Desktop File Manager Icon Gnome le a O Bhs Main Menu Task Buttons Workspace Area Button The Gnome Main menu provides access to applications Click the Main Menu button shaped like a footprint in the Gnome panel to open the Main menu Point to an item on the menu to display that item s menu Figure 43 The Gnome Main menu 4 Programs amp Favorites Applets KDE menus PH Run Lock screen Log out Tip An easy way to set up your desktop so that you can toggle between the arrayWoRx software and a Windows application is to run arrayWoRx on one workspace and run Windows through VMWare on another ppliedPrecision Chapter 10 Introducing Linux and Gnome 111 Working with Windows This section describes a few techniques for opening moving resizing and clo
156. you can see the spot and center it i Drag the zoom wheel down to zoom in until the spot takes up about a quarter of the Image window ii Click PS i Click a point near the middle of the spot 5 Zoom in until the spot fills the image window Right click on an area or click the down zoom arrow to incrementally zoom in Middle click on an area or click the up zoom arrow to zoom out 04 720046 000 Rev D 0804 72 arrayWoRx Standard amp Auto User s Manual 6 Pan to adjust the spot s position in the window 1 Click 7 The pointer turns into a hand shaped icon ii Drag the spot to move it 7 Define the ROI i From the TOOLS menu choose DATA INSPECTOR to open the Data Inspector ii Click Select Box ili In the Image window point to the top left corner of a square that would enclose the spot iv Drag the bottom right corner of the ROI box toward the bottom right corner of the Image window to enclose the spot with the ROI box v Drag the ROI box to move it over the spot vi To make the ROI a circle instead of a square select Constrain to Circle ppliedPrecision CONTENTS File Menu Scan Menu Utilities Menu Help Menu Buttons Slide View Scan Settings Preset Scan Settings Tabs Panel View Scan Table The Status Bar 7 MAIN SCAN WINDOW REFERENCE This chapter briefly describes the features of the Main Scan window If you are looking for instructions on the use of a particular feature check
157. your warranty Third party Windows applications are allowed Configuration subject to change without notice ppliedPrecision Chapter 2 System Overview 11 Installation Requirements and Recommendations Electrical Requirements Voltage 100 240 VAC 10 Frequency 50 60 Hz Circuit Rating 5 A maximum 6 3 A fuse Transient over voltages in accordance with Installation Category II in IEC 664 Space Requirements 4 ft x 2 5 ft 120 cm x 80 cm of counter space Operating Environment Requirements Indoor use Temperature 65 77 F 18 25 C daily variation of no more than 3 F 1 8 C Humidity maximum relative humidity less than or equal to 50 High humidity can result in condensation on the camera window Excessive humidity may also reduce filter life Altitude up to 6550 ft 2000 m POLLUTION DEGREE 2 in accordance with IEC 664 Communication Recommendations It is suggested that the workstation be connected to a local area network for data storage A connection to the Internet will provide access to Applied Precision s web site To connect the system to a network you will need an IP address domain name server address and network mask Important Applied Precision LLC is not responsible for damage or harm to the workstation or scanner due to network security breaches Having a telephone in the same room as the system is also recommended to facilitate communication with Technical Sup

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