ALT•AST (GPT•GOT)
Contents
1. 402 12780 qxd 8 3 04 6 02 PM Page 1 Cholestech LDX ALTeAST GPT GOT Alanine Aminotransferase Glutamate Pyruvate Transaminase and Aspartate Aminotransferase Glutamate Oxaloacetate Transaminase Test Cassette REF 12 788 CLIA WAIVED These tests are waived under CLIA 88 regulations If a laboratory modifies the test system instructions then the test is considered highly complex and subject to all CLIA requirements For professional in vitro diagnostic use U CHOLESTECH ss 402 12780 3347 Investment Blvd Hayward CA 94545 U S A Tel 800 733 0404 Fax 510 732 7227 www cholestech com Cholestech LDX is a registered trademark of Cholestech Corporation MiniPet is a trademark of TriContinent Scientific Inc 2004 Cholestech Corporation 402 12780 REVA One or more of the following patents may apply U S Patents 4 477 575 4 816 224 and 5 110 724 Authorized Representative AR MED Ltd Runnymede Malthouse Egham TW20 9BD United Kingdom Refer to the CD in the analyzer package for instructions in English The instructions are available from your local distributor Le CD contenu dans l emballage de l analyseur inclut les directives d utilisation en fran ais Le mode d emploi est disponible aupr s du distributeur local Anweisungen auf Deutsch befinden sich auf der CD in der Verpackung des Analysegerats Die Anleitung ist von Ihrem Handler erh ltlich Fare riferimento al CD nella confezione dell anal2. Let cassette sit at room temperature for 10 minutes 2 Remove the cassette from its pouch Hold the cassette by the short side only Do not touch the black bar or the brown magnetic stripe Put the cassette on a flat Surface NOTE Gloves should be worn when working with blood samples 3 Press RUN In a few seconds the screen will read 4 The cassette drawer will open The screen will read 5 Place the sample into the cassette well Use a Cholestech LDX Capillary Tube for fingerstick samples Use the MiniPet Pipette for controls or venous blood samples NOTE Fingerstick samples must be applied within five 5 minutes or the blood will clot 6 Keep the cassette flat after the sample has been applied Place the cassette into the drawer of the Analyzer immediately The black bar must face the Analyzer The brown stripe must be on the right 7 Press RUN The drawer will close During the test the screen will read 8 Put everything that touched the blood sample or control in a biohazard waste container 9 When the test is complete the Analyzer will beep The screen will read 10 Press DATA to show more results 11 When the results are outside the measuring range the screen will read 12 If there is a problem with the test a message will appear on the screen See the Troubleshooting section of the Cholestech LDX System User Manual if this happens 13 When the drawer opens remove the cassett
3. 5 10 349 Serum 52 0 914 0 5 0 971 19 383 Fingerstick 24 0 921 4 3 0 931 15 65 AST measured using the Cholestech LDX cassette was compared with a validated reference traceable to the IFCC reference method Asparate Aminotransferase Results X Reference Method serum Y Cholestech LDX Analyzer Sample No of y Correlation Range Type LDX Pairs Slope Intercept Coefficient of Values Serum 109 0 97 1 6 0 983 12 396 Accuracy Sample Type The results from venous whole blood and fingersticks were compared to the serum values obtained on the LDX Results X Serum on the LDX Y Whole Blood or Fingerstick on the LDX Sample No of y Correlation Range Type LDX Pairs Slope Intercept Coefficient of Values Venous Whole Blood 46 1 08 0 3 0 998 13 343 Fingerstick 21 0 86 44 0 934 13 65 ALT WAIVER DATA A consumer accuracy study was done with 60 lay untrained people who reported no medical or laboratory experience These people were asked to perform one ALT test each by following the directions in the package insert The samples used for testing were 60 whole blood samples that were either native samples or samples spiked with ALT A total of 3 professional trained people also assayed the 60 samples 20 samples per trained person in order to obtain comparative data between trained and untrained populations The results were analyzed by Deming regression statistics as described below ALT Consumer Accuracy Study trained x
4. ansferase catalyzes the transfer of amino groups from L alanine to alpha ketoglutarate producing pyruvate and glutamate Pyruvate oxidase in the presence of oxygen oxidizes the pyruvate to acetylohosphate and hydrogen peroxide In a reaction catalyzed by horseradish peroxidase the peroxide reacts with an indicator dye to form a blue color at a rate proportional to the ALT concentration of the sample The resultant color in the reaction is measured by reflectance photometry Alanine Aminotransferase L Aspartic acid alpha Ketoglutarate _ ______ _ _ gt Pyruvate Oxidase Pyruvate Phosphate O2 H20 _ gt Acetylphosphate COs H202 Peroxidase SE ae Pyruvate Glutamate Indicator H202 Colored Blue Dye H20 The Cholestech LDX Analyzer measures aspartate aminotransferase by an enzymatic method based on the method formulation of Katsuyama et al 2 Aspartic acid aminotransferase catalyzes the transfer of amino groups from L Aspartic acid to alpha Ketoglutarate producing oxaloacetate and glutamate Oxaloacetate Decarboxylase converts the Oxaloacetate to Pyruvate by the removal of CO Pyruvate oxidase in the presence of oxygen oxidizes the pyruvate to acetylphosphate and hydrogen peroxide In a reaction catalyzed by horseradish peroxidase the peroxide reacts with an indicator dye to form a blue color at a rate proportional to the AST concentration of the sample The resultant color
5. ation Program recommendations for measurement of low density lipoprotein cholesterol executive summary Clin Chem 1995 41 1414 20 Tietz NW ed Fundamentals of Clinical Chemistry Philadelphia Pa WB Saunders Co 1987 Siedel J Hagele EO Ziegenhorn J Wahlefeld AW Reagent for the enzymatic determination of serum total cholesterol with improved lipolytic efficiency Clin Chem 1983 29 1075 80 i w P E In vitro diagnostic medical device JJ mT m Catalog number Attention See instructions for use Single use Do not use if package is damaged or open Lot number Use by Temperature K E 8 p
6. axis vs untrained y axis Slope 95 y y Intercept 95 n Slope Confidence Interval Intercept Confidence Interval r 60 0 976 0 934 to 1 019 1 0 1 4 to 3 3 0 996 AST WAIVER DATA An untrained user study was conducted in which participants were given only the test instructions and asked to perform testing of 3 blinded randomized samples The samples consisted of serum pools prepared at three levels The participants were not given any training on the use of the test A total of 72 participants were enrolled from three sites representing a diverse demographic educational age gender etc population The table below presents the summary of the performance Level 1 Level 2 Level 3 N 72 71 72 Mean 52 9 U L 187 4 U L 289 8 U L CV 7 4 4 4 5 1 Observed 244 346 170 211 244 346 Range Percent of 97 2 100 98 6 Results in the 70 72 71 71 71 72 Range 15 95 Cl 90 3 to 99 7 95 Cl 94 9 to 100 95 Cl 97 5 to 100 REFERENCES 1 Expert Panel on Detection Evaluation and Treatment of High Cholesterol in Adults Executive summary of the Third Report of the National Cholesterol Education Program NCEP Expert Panel on Detection Evaluation and Treatment of High Cholesterol in Adults Adult Treatment Panel Ill JAMA 2001 285 2486 97 Bachorik PS Ross JW for the National Cholesterol Education Program Working Group on Lipoprotein Measurement National Cholesterol Educ
7. e Put it in a biohazard waste container Leave the Analyzer drawer empty when not in use 14 Record the results 15 To run another cassette press RUN The screen will read 16 Repeat the test procedure 17 Otherwise after 4 minutes a beep will sound and the screen will read If the RUN button is not pushed within 15 seconds the drawer will close Then the screen will go blank QUALITY CONTROL Quality control should be run routinely to show that your system is giving accurate results We recommend the following quality control procedures for the Cholestech LDX System Choice of Materials A high and a low control for each analyte is preferred Cholestech recommended controls work well with the Cholestech LDX System If you use other controls you will need to set ranges for the Cholestech LDX System Handling e Follow the instructions that come with your controls e Check the expiration date before use Do not use if expired e See Running a Test for procedure External Quality Control External controls must also be used to demonstrate that the reagents and the assay procedure perform properly Liquid Level 1 and Level 2 Controls are available from Cholestech Controls must be tested e With each new shipment of cassettes even if cassettes are from the same lot previously received e With each new lot of cassettes e As otherwise required by your laboratory s standard quality control procedu
8. ested on samples from newborns or pediatric patients Hemolysis breakdown of the red blood cells should be avoided as this will increase the level of ALT and AST and cause inaccurate results e Samples with ALT or AST enzyme activity greater than 1000 U L may consume the substrate prior to the measurement of enzyme activity and could yield falsely low results Some substances may cause false results with enzymatic tests The substances listed below were tested for interference with the ALT AST test Less than 10 interference was seen at the levels shown Substance Concentration mg dL Hemoglobin 75 Gemfibrozil Lopid 15 Bilirubin 5 Oxytetracycline 4 Ditaurobilirubin 5 Probucol Lorelco 32 5 Ascorbic Acid 1 Nicotinic Acid Niacin 10 Urea 500 Clofibrate Atromid 80 Uric Acid 15 Lovastatin Mevacor 4 Creatinine 30 L Dopa Levodopa 0 2 Glutathione 1 Cimetidine Tagamet 5 Lactate 100 Nitrofurantoin 1 Fructose 30 Gentisic Acid 0 5 Lactose 100 Methyl dopa 0 2 Cysteine 7 Sulfamethoxyzole 16 402 12780 qxd 8 3 04 6 02 PM Page 2 Glucose 1200 Pravastatin Pravachol 0 8 Pyruvic acid 0 2 Simvastin Zocor 4 0 Triglycerides 450 Fluvastatin Lescol 4 0 Hematocrits between 30 and 50 do not affect results EXPECTED VALUES Alanine Aminotransferase Reference Interval 99 F 37 C 10 40 U LS Aspartate Aminotransferase Reference Interval 99 F 37 C 10 30 U L3 PERFORMANCE CHARACTERISTICS Precision A st
9. in the reaction is measured by reflectance photometry Aspartate Aminotransferase L Aspartic acid alpha Ketoglutarate ____________ gt _ Oxaloacetate Glutamate Oxaloacetate Decarboxylase Oxaloacetate Pyruvate Oxidase Pyruvate Phosphate O2 H20 _ gt Acetylphosphate CO2 H202 Peroxidase eae Pyruvate CO Indicator H202 Colored Blue Dye H20 A brown magnetic stripe on each cassette contains the calibration information required for the Cholestech LDX Analyzer to convert the reflectance reading to the ALT and AST concentration in U L 37 C 99 F for whole blood The Lot number and Expiration date are also contained on the stripe ALANINE AMINOTRANSFERASE AND ASPARTATE AMINOTRANSFERASE CASSETTE Each ALTeAST Cassette contains ALT AST L Alanine ug 91 9 L Aspartic Acid ug 67 1 alpha Ketoglutaric Acid ug 6 4 6 4 Sodium phosphate monobasic ug 9 6 8 1 Oxaloacetate Decarboxylase U 0 34 Pyruvate Oxidase Aerococcus viridans U 2 4 2 0 Peroxidase horseradish U 3 6 3 0 Ascorbate Oxidase Cucurbita sp U 1 7 1 7 Indicator 2 3 5 di tert butyl 4 hydroxyphenyl 4 5 bis 4 dimethylaminophenyl imidazole ug 6 1 6 1 Nonreactive ingredients Buffers and stabilizers Cassette Storage and Stability Cassettes must be stored in the sealed foil pouches Cassettes may be used until the date printed on the pouch when stored in a refrigerator 36 46 F 2 8 C O
10. it may cause poor results Use a MiniPet Pipette and tip to place blood into the cassette Whole blood should be used within 30 minutes Samples should be at room temperature for testing Mix all samples by inverting gently 7 8 times before testing PRECAUTION All blood samples and containers capillary tubes and materials that have come in contact with blood should be handled as if capable of transmitting infectious disease and discarded into a biohazard waste container after use Calibration No calibration is done by the user Test information is on the brown stripe of the cassette The brown stripe is read by the Cholestech LDX each time a cassette is run An optics check should be run on the Analyzer each day that patient samples are tested See the Cholestech LDX System User Manual for instructions TEST PROCEDURE Materials Provided Test Cassettes Additional Materials Required Cholestech LDX Analyzer and power supply Alcohol swabs and gauze for cleaning puncture site Lancets for capillary blood collection Cholestech LDX Capillary Tubes with lithium heparin anticoagulant Cholestech LDX Capillary Plungers Gloves Biohazard waste containers Quality control material MiniPet Pipette and tips or micropipetter that will deliver 35 uL for use with venipuncture samples and quality control material e Vacuum collection tubes needles and tube holders if sample is to be collected by venipuncture Running a Test 1
11. izzatore per istruzioni in italiano Le istruzioni sono disponibili presso il distributore di zona Consulte el CD incluido en el envase del analizador para obtener instrucciones en espanol Tambi n puede pedir las instrucciones a su distribuidor local Consulte o CD no pacote do analisador para instru es em Portugu s As instru es est o dispon veis junto do seu distribuidor local Der henvises til den vedlagte CD i analysatorpakken for instruktioner pa dansk Instruktionerne fas hos den lokale forhandler Se CD n i analysatorforpackningen betr ffande instruktioner pa svenska Instruktionerna finns att fa hos din lokala terf rs ljare Avatp Ete oto CD otn ouokeuacia tou avaAuTH yra o ny es ota EAAnViKa OL odnyies eivat SLtaGEoLlMES ATO TOV TOTILKO SLAVOLEA OAs INTENDED USE An in vitro diagnostic for the quantitative determination of alanine aminotransferase ALT and aspartate aminotransferase AST in whole blood INTRODUCTION Alanine aminotransferase is an enzyme that catalyzes the conversion of alanine to pyruvate Aspartic acid aminotransferase is an enzyme that catalyzes the conversion of aspartic acid to oxaloacetate Both are found in cardiac and skeletal muscle the liver and other tissues ALT is most prevalent in the liver ALT and AST levels are a reflection of alterations in liver function and therefore are a valuable measurement of damage to the liver Liver damage may be due to chronic alcohol or drug i
12. ngestion or infection There are a number of lipid lowering drugs available to treat hyperlipidemia A side effect of such therapy can be a persistent increase in serum ALT and or AST to more than 3 times the upper limit of normal in about 1 of patients receiving lipid lowering therapy It is suggested that patients undergoing lipid lowering drug therapy should be tested for ALT and AST before baseline and shortly after initiation of therapy and then periodically thereafter to determine the ALT and AST levels Alanine aminotransferase and aspartic acid aminotransferase can be measured from a single drop of blood using the Cholestech LDX System s rapid accurate technology An AST ALT ratio is calculated using the measured values SUMMARY AND EXPLANATION The Cholestech LDX System combines enzymatic methodology and solid phase technology to measure ALT and AST Samples used for testing can be whole blood from a fingerstick collected in a heparin coated capillary tube or venous whole blood The sample is applied to a Cholestech LDX ALT AST cassette The cassette is then placed into the Cholestech LDX Analyzer where a unique system on the cassette separates the plasma from the blood cells The plasma flows to both sides of the cassette and is transferred to the ALT and AST reaction pads The Cholestech LDX Analyzer measures alanine aminotransferase by an enzymatic method based on the method formulation of Katsuyama et al 2 Alanine aminotr
13. r the cassettes may be stored for up to 30 days at room temperature less than 86 F 30 C e Once cassettes have been stored at room temperature they should not be returned to the refrigerator e Do not use a cassette beyond the printed expiration date e Do not use a cassette that has been stored at room temperature for more than 30 days Do not reuse cassettes Cassette Handling Cassettes should sit at room temperature for 10 minutes before opening the pouch Use the cassette as soon as the pouch is opened Sample Type Please Note The Cholestech LDX System is CLIA waived for fingerstick or venous whole blood unprocessed samples only If you run serum or plasma on the Cholestech LDX you will be classified as moderately complex and will have to comply with the regulations for moderate complexity See the Cholestech LDX System User Manual for a summary of these regulations Sample Handling e Sample Volume 35 uL of whole blood Fingerstick whole blood e Collect the sample from a fingerstick into a Cholestech LDX Capillary Tube See the Fingerstick Procedure in the Cholestech LDX System User Manual e Place the blood into the cassette within 5 minutes of collection e Blood from the fingerstick should flow freely Too much squeezing of the finger may Cause poor results Venous whole blood e Collect blood into a green top tube heparin anticoagulant NOTE Do not use a tube with any other additives because
14. res e If you are not running the Cholestech LDX under CLIA waived status or if your local or state regulations require more frequent testing of quality control material then quality control must be performed in compliance with those regulations Good Laboratory Practice principles suggest that external controls must be run whenever the laboratory director has any question about test system integrity or operator technique e g when reagents may have been stored or handled in a way that can degrade their performance or when operators have not performed a particular test in recent weeks If the controls do not perform as expected repeat the test or contact Cholestech Technical Service before testing patient samples The quality control results must be in range before testing patient samples See the Cholestech LDX System User Manual if they are not Please call Cholestech Technical Service at 800 733 0404 if you have any questions about quality control RESULTS ALT and AST test results will show on the screen when the test is complete Calculated results are shown after the DATA button is pressed LIMITATIONS e The measuring range for ALT is 10 400 U L 99 F 37 C Results outside this range will appear as lt 10 U L or gt 400 U L e The measuring range for AST is 10 400 U L 99 F 37 C Results outside this range will appear as lt 10 U L or gt 400 U L e Performance of the Cholestech LDX System has not been t
15. udy was conducted according to NCCLS protocol EP5 A Evaluation of Precision Performance of Clinical Chemistry Devices Approved Guideline 1998 4 Alanine Aminotransferase Commercial Commercial Control Material Control Material Frozen Level 1 Level 2 Serum Pool X U L 31 58 169 Within run CV 3 2 3 1 3 4 Total CV 5 4 4 6 6 5 Whole Blood Within Run Precision X U L 55 SD U L 2 3 CV 4 2 Aspartate Aminotransferase Commercial Commercial Control Material Control Material Frozen Level 1 Level 2 Serum Pool X U L 31 106 277 Within run CV 6 1 3 5 3 8 Total CV 8 8 4 4 5 2 Whole Blood Within Run Precision X U L 58 SD U L 2 8 CV 4 8 ACCURACY METHOD COMPARISON ALT measured using the Cholestech LDX cassette was compared with a validated reference method traceable to the IFCC reference method and with a bench top point of care chemistry analyzer Alanine Aminotransferase Results X Reference Method serum Y Cholestech LDX Analyzer Sample No of y Correlation Range Type LDX Pairs Slope Intercept Coefficient of Values Venous Whole Blood 53 1 001 0 3 0 966 10 349 Serum 54 1 007 2 8 0 960 14 391 Fingerstick 24 1 013 3 5 0 930 15 65 Results X Bench Top Point of Care Chemistry Analyzer Y Cholestech LDX Analyzer Sample No of y Correlation Range Type LDX Pairs Slope Intercept Coefficient of Values Venous Whole Blood 53 0 916 0 3 0 97
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