TnI Troponin l
Contents
1. 0 010 ng mL r 0 96 StatusFirst Tnl serum 1 197 x StatusFirst Tni Plasma EDTA r 0 98 CLINICAL PERFORMANCE A total of 228 patients whole blood and plasma specimens collected in EDTA and or lithium heparin tubes were tested TnI concentrations in plasma samples from lithium heparin tubes were determined by both Access AccuTnI and the StatusFirst Tnl test Tni concentrations in whole blood samples from EDTA tubes were determined by both Triage Cardiac Panel and StatusFirst Tnl test Tni levels in plasma from EDTA tubes and in whole blood from lithium heparin tubes were tested on the StatusFirst Tnl test only and used to determine the correlation of different matrices and anticoagulants The sensitivity and specificity of the StatusFirst Tnl test was evaluated against Access AccuTnI and the StatusFirst Tnl performance was correlated to Access AccuTnI or Triage Cardiac Panel Page 11 Clinical Sensitivity and Specificity The StatusFirst Tnl test results in this clinical study were analyzed using the Receiver Operator Characteristics ROC curve The diagnostic utility of the StatusFirst Tnl test is demonstrated by the area under the ROC curve of gt 0 99 which indicates that the StatusFirst Tnl test is as effective as Access AccuTnI in the diagnosis of AMI Figure 1 and Table 5 Figure 1 ROC curve comparing with Access AccuTnI a Source of the Curve 0 84 Stat2. Self Check items should pass before testing patient samples The operator may then proceed to Calibration QC If the DXpress Reader is left on or in power save mode the operator should perform Self Check daily as follows 1 From the Main Menu select 2 RUN QC 2 Select 1 SELF CHECK 3 Self Check takes about 15 seconds PASS or FAIL results will be displayed printed when testing is completed Page 4 Calibration QC Each day of patient testing use the Calibration Set see DXpress Reader manual to ensure the DXpress Reader functions properly From the Main Menu select 2 RUN QC Select 2 CALIBRATION QC Scan or manually enter the Operator ID barcode Scan the Calibrator ID barcode found on the back of the Calibrator Insert the Calibrator into the reader be sure to close the Tray and press ENTER Follow prompts displayed on the screen 6 Calibration QC result will be displayed printed when the calibration is completed Calibration QC should pass before running daily patient testing 7 Blank Calibration is not necessary for TnI product Select CANCEL and press ENTER ahwonh External QC Perform External QC once for each new lot of StatusFirst Tnl test devices 1 From Main Menu select 2 RUN QC 2 Select 3 EXTERNAL QC 3 Using external controls follow the same procedure as if running a patient sample see section Testing Patient Sample Procedure below The onl
3. Warnings For in vitro diagnostic use only Carefully follow the instructions for use Wear disposable gloves while handling patient samples Patient samples used test devices and transfer pipettes should be treated as if potentially infectious and should be discarded as biohazard materials according to local regulations Thoroughly wash hands afterwards and observe the appropriate regulations procedures for disposal of all used materials samples test devices and transfer pipettes The result obtained from the StatusFirst Tnl test device does not provide a definitive diagnosis and should be interpreted by the physician in conjunction with other laboratory test results and patient clinical findings Avoid cross contamination of samples by using a new transfer pipette for each sample Keep the test device in the sealed pouch until ready for use Do not use the test device if the pouch is damaged or the seal is broken Do not use the test device after the expiration date printed on the pouch This is a quantitative test and therefore no visual interpretation of the result should be made The test must be read at 15 minutes after sample addition to ensure an accurate result Storage and Stability Store the StatusFirst Tnl test device between 2 and 8 C 35 to 46 F until the expiration date printed on the pouch is reached The StatusFirst Tnl test device in its sealed pouch is stable at room temperature 18 to 30 C 6
4. for the calibration as designated by the American Association for Clinical Chemistry StatusFirst Tnl Lithium Heparin Plasma 0 323 x AccuTnI Plasma Lithium Heparin 0 007 ng mL r 0 93 with n 182 StatusFirst Tnl Whole blood EDTA 0 623 x Triage Whole blood EDTA 0 065 ng mL 0 92 with n 95 Page 13 REFERENCES ie World Health Organization Report of the Joint Internaltional Society and Federation of Cardiology World Health Organization Task Force on Standardization of Clinical Nomenclature Nomenclature and criteria for diagnosis of ischemic heart disease Circulation 1979 59 607 9 Veterans Health Administration Ischemic heart disease inclusive of JCAHO AMI Veterans Health Administration Report 2005 Mar NQMC 000931 Heidenreich P A Go A Melsop K A et al Prediction of risk for patients with unstable angina AHRQ Publication No 01 E001 December 2000 Number 31 Mehegan J P amp Tobacman L S Cooperative interaction between troponin molecules bound to the cardiac thin filament J Biol Chem 266 2 966 972 1991 Ebashi S Ca2 and the contractile proteins J Mol Cell Cardiol 16 2 129 136 1984 Bodor GS et al Development of monoclonal antibodies for an assay of cardiac troponin and preliminary results in suspected cases of myocardial infarction Clin Chem 38 11 2203 2214 1992 Takahashi M et al Use of immunoassay for measurement of skeletal troponin
5. utilizing isoform specific monoclonal antibodies Clin Biochem 29 4 301 308 1996 Adams J E et al Biochemical markers of myocardial injury Is MB creatin kinase the choice for the 1990s Circulation 88 750 763 1993 Apple F S et al Cardiac troponin CK MB and myoglobin for the early detection of acute myocardial infarction and monitoring of reperfusion following thrombolytic therapy Clinica Chimica Acta 237 1 2 59 66 1995 Adams J E et al Improved detection of cardiac contusion with cardiac troponin American Heart J 131 2 308 312 1996 Adams J E et al Diagnosis of perioperative myocardial infarction with measurement of cardiac troponin I N Eng J Med 330 10 670 674 1994 Machler H et al Preoperative myocardial cell damage in patients with unstable angina undergoing coronary artery bypass graft surgery Anesthesiology 81 6 1317 1320 1994 Hossein Nia M et al Cardiac troponin release in heart transplantation Ann Thorac Surg 61 1 277 278 1996 Wu A et al Prognostic value of cardiac troponin in patients with chest pain Clin Chem 42 4 651 652 1996 Braunwald E et al ACC AHA 2002 guideline update for the management of patients with unstable angina and non ST segment elevation myocardial infarction ACC AHA Practice Guidelines ACC Bethesda MD amp AHA 2002 Wu HBA Apple PS Gibler B Jesse RL et al National Academy of Clinical B
6. 4 to 86 F for 15 days provided the expiration date printed on the pouch is not exceeded Page 3 Sample Collection and Preparation e The StatusFirst Tnl test device is to be run using whole blood serum or plasma samples collected using EDTA or lithium heparin as the anticoagulant e Whole blood samples must be tested within 4 hours and plasma or serum samples must be tested within 8 hours of collection e Whole blood serum or plasma samples should be stored and or transported in refrigerated conditions e If longer storage is required plasma or serum samples should be kept frozen at 20 C 4 F or lower e Allow patient samples to equilibrate to room temperature prior to testing e Grossly hemolyzed samples should not be used e Mix the blood specimen gently inverting the tube several times before using e For frozen samples thaw completely and mix thoroughly before use PROCEDURE DXpress Reader a Consult the DXpress Reader User Manual For DXpress Reader installation startup and complete instructions refer to the DXpress Reader User Manual The operator must consult the DXpress Reader User Manual prior to use and become familiar with the processes and quality control procedures QC Procedure Self Check Each time the DXpress Reader is turned on Self Check is automatically performed Self Check takes about 15 seconds PASS or FAIL results will be displayed printed when testing is completed All
7. Catalog number Tn For the Rapid Quantitative Determination of Troponin I in human Whole Blood Plasma or Serum w StatusFirst 60501 Manufacturer Manufactured for Princeton BioMeditech Corporation LifeSign LLC 4242 US Route 1 71 Veronica Avenue Monmouth Junction NJ 08852 Somerset NJ 08873 www pbmc com Tel 1 800 526 2125 www lifesignmed com Authorized Representative MT Promedt Consulting GmbH Altenhofstrasse 80 66386 St Ingbert Germany 49 68 94 58 10 20 P 56611 Version 226 10 24 06 StatusFirst Tnl Explanation of Symbols CE F opm 6 lt s v o Eg CE Marking of Conformity Manufacturer Catalog number Expiry date Use by Lot number In Vitro Diagnostic medical device Consult instructions for use Store between 2 C and 8 C Manufactured for m e pe D o gt Cc gt e N D a D D gej D n D gt o a lt D Contents Test Device Data Chip Package Insert Desiccant Transfer Pipette Intended Use For the quantitative determination of Troponin as an aid in the diagnosis of myocardial infarction and acute coronary syndrome Contains sufficient for lt n gt tests Do not reuse StatusFirst Tnl For in vitro diagnostic use Intended Use StatusFirst Tnl Troponin is a rapid test for in vitro quantitative determination of Troponin in human whole blood plasma or serum The device is intended for use wi
8. TEMI patients Therefore measurement of sensitive and specific cardiac protein markers in the blood provides an extremely useful differential diagnostic tool enabling prompt management of patients presenting with symptoms of AMI Troponin and troponin T are two specific contractile proteins of the cardiac muscle regulatory complex that have gained popularity as cardiac specific markers for AMI Three subunits of the troponin complex I T C are bound with tropomyosin to actin in the thin filament of the myofibril Cardiac troponin cTnl prevents muscular contraction and is exclusively present in the cardiac muscle The Troponin complex together with tropomyosin forms the main component that regulates the Ca2 sensitive ATPase activity of actomyosin in striated muscle skeletal and cardiac Different isoforms of Tnl exist in the skeletal and cardiac muscles sTnl and cTnl respectively with distinct structural heterogeneity between these isoforms that allow Page 1 the production of isoform specific antibodies Specifically the amino terminal amino acid sequence of cTnl has 31 amino acid residues that are not present in sTnl The utility of determining the serum levels of the different isoforms of TnI has been studied Detection of cTnl in the serum has been investigated as an aid in the determination of myocardial damage in patients with AMI Determining the serum level of cTnl provides diagnostic value in identifying patien
9. device utilizes biotin coupled anti Tnl antibody streptavidin solid phase chromatographic immunoassay technology to quantitatively determine the concentration of TnI in human whole blood plasma and serum specimens After a sample has been dispensed into the sample well the StatusFirst Tnl test device is placed in the DXpress Reader The DXpress Reader displays the TnI concentration 15 minutes after sample addition The DXpress Reader is programmed to convert the intensity of the test band as indicated by the TnI line on the test device into a concentration of Tn automatically by using lot specific calibration factors supplied with each box of test devices The TnI concentration in the sample correlates with the intensity of the test band Reagents The StatusFirst Tnl test device contains all required reagents including dye conjugated monoclonal anti Tnl antibody biotin conjugated polyclonal anti Tnl antibody and streptavidin immobilized at the test band No other reagents are required Page 2 Materials Provided Each box contains the following 20 StatusFirst Tnl test devices each individually sealed in a foil pouch with a desiccant and a single use transfer pipette 1 lot specific Data Chip with calibration information 1 package insert Materials Equipment Required But Not Provided 1 DXpress Reader part no LSR 2000 2 Commercially available TnI Controls for external Quality Control QC Precautions and
10. iochemistry Standards of Laboratory Practice Recommendations for the use of cardiac markers in coronary artery disease Clin Chem 1999 45 7 1104 1121 Manufactured by PBM Princeton BioMeditech Corporation 4242 US Route 1 Monmouth Junction NJ 08852 U S A Tel 1 732 274 1000 Fax 1 732 274 1010 www pbmc com
11. ite QUALITY CONTROL External Controls Good laboratory practice includes the use of external controls to ensure proper test device performance It is recommended that prior to using a new lot or shipment of StatusFirst Tnl test devices the performance of the lot be confirmed by testing with external controls see section Materials Equipment Required But Not Provided to ensure the test devices will deliver the correct test result The frequency of QC testing should be determined according to individual laboratory standard QC procedures Upon confirmation of the expected results the test devices are ready for use with patient samples If external controls do not perform as expected do not use the test devices and contact LifeSign Technical Services at 1 800 526 2125 Internal Controls StatusFirst Tnl test device has a built in control that satisfies the requirements of testing a control on a daily basis The control line is an internal positive procedural control A distinct reddish purple control line should appear at the control position if the test is performed properly an adequate sample volume is used the sample and reagent are wicking on the membrane and the reagents at the control line are reacting with the conjugate color indicator In addition a clear background may be considered a negative procedural control If the test is performed correctly and the device is working properly the background in the result window will beco
12. maximum concentration of substance indicated Table 3 No substance demonstrated significant cross reactivity i e all cross reactivities lt 0 05 when added to sample containing a TnI concentration of approximately 0 5 0 7 ng mL Table 3 Cross reactivity study Protein Peptides Actual Conc ug mL Cross reactivity Tropomyosin cTnT cTnC cMyosin light chain Myoglobin CK MB BNP NT proBNP Page 9 Drugs Sixty drugs were assessed for potential interference in the StatusFirst Tnl test device Table 4 The list of drugs encompassed common prescription and over the counter compounds as well as medications often prescribed in a cardiac related patient population The drugs were tested at concentrations as recommended in the CLSI Approved Guideline EP7 A Interference Testing in Clinical Chemistry or at least three times the highest concentration reported following a therapeutic dosage No significant interference with the StatusFirst Tnl measurement was observed for the drugs listed in the table below Table 4 List of drugs tested for the interference study Acetaminophen Dipyridamole Oxazepam Actetylsalicylic acid Aspirin Dopamine Oxytetracycline Allopurinol Enalapril maleate Phentobarbital Alteplase Erythromycin Phenytoin Ambroxol Fluvastatin Lescol Pravastatin Amiodarone Furosemide Probenecid Amlodipine Besylate Glybu
13. me clear and provide a distinct result The DXpress Reader will report Control Valid and the test result for TnI concentration in ng mL unit when Internal Control QC is satisfied LIMITATIONS e The results of the StatusFirst Tnl test should be used in conjunction with the total Clinical presentation of the patient and other laboratory information available Medical decisions should not be based on a single point in time test result of the StatusFirst Tni test The published guideline recommends collecting at least three blood samples during the early triage period e Other substances and or factors not listed e g technical or procedural errors may interfere with the test and cause false results Page 7 PERFORMANCE CHARACTERISTICS Limit of Detection The limit of detection LoD represents the lowest concentration of Tnl that can be reliably differentiated from zero The LoD of 0 05 ng mL was determined according to Clinical and Laboratory Standards Institute CLSI guideline EP17 A Limit of Quantitation The limit of quantitation LoQ is the lowest TnI concentration that can be reproducibly measured with a total coefficient of variation of at most 20 The LoQ was determined to be 0 1 ng mL Linearity Human cardiac troponin complex from the National Institute of Standards amp Technology NIST was spiked into human blood to a final concentration of 32 ng mL cardiac troponin I and then plasma was separated from
14. pense into the sample well When drawing sample into the transfer pipette avoid introducing air bubbles Do not touch the sample well or test device with the tip of the transfer pipette Fill sample up to line e Press ENTER e Insert the test device in the Reader tray close the Reader tray e After 15 minutes of incubation the DXpress Reader will automatically display the results on the screen e Results may be printed by pressing the PRINT button e At this point the test device may be removed and appropriately discarded Report and Interpretation of Results e The range of TnI concentrations reported by the test device system is 0 05 ng mL to 30 ng mL Results below or above this range will be shown as lt 0 05 ng mL or gt 30 ng mL respectively e Valid results will be displayed on the Reader as Control Valid Tol xx xx ng mL e Invalid results If the sample fails to migrate properly or the reagents fail the Reader will display Control Invalid Tni Invalid In this case repeat the test with a new test device Page 6 Recommended Decision Threshold Values The clinical cutoff of StatusFirst TnI was determined as 0 2 ng mL TnI when compared to Access AccuTnI t and Triage Cardiac Panel t However laboratories should establish their own diagnostic cutoff concentration based on the clinical practice at their respective institutions tRegistered trademark of Beckman Coulter Registered trademark of Bios
15. ride Procainamide Ampicillin Heparin Propranolol Ascorbic acid vitamin C Hydralazine Quinidine Atenolol Hydrochlorothiazide Spironocactone Atorvastatin calcium Indomethacin Sulfamethoxazole Caffeine Isosorbide dinitrate Theophylline Captopril Lisinopril L thyroxine Chloramphenicol Methaqualone Trimethoprim Chlordiazepoxide Methyl DOPA Verapamil Cinnarizine Milrinone lactate Warfarin Clopidogrel bisulphate Nicotine Cyclosporine A Nifedipine Diclofenac Nitrofurantoin Digitoxin Nitroglycerin Digoxin Noraminopyren Dipyrone Diltiazem Nystatine Page 10 Other Potentially Interfering Substances When added to a sample containing TnI hemoglobin up to 0 5 g dL bilirubin up to 20 mg dL triglycerides up to 1 g dL and human albumin up to 16 g dL did not interfere with the recovery of Tnl Hook Effect No high dose hook effect was observed for TnI concentrations up to 500 ng mL Blood Plasma and Serum Correlation Blood plasma and serum comparison study was performed with clinical samples A comparison analysis between blood plasma and serum showed the following correlations StatusFirst Tni EDTA Plasma 0 958 x StatusFirst Tni EDTA Whole blood EDTA 0 002 ng mL r 0 95 StatusFirst Tnl Lithium Heparin Plasma 0 946 x StatusFirst Tni Whole blood Lithium Heparin
16. th the DXpress Reader to provide quantitative results as an aid in the diagnosis of myocardial infarction and acute coronary syndrome ACS Summary and Explanation Acute coronary syndrome ACS is the leading cause of morbidity and mortality among both men and women affecting more than 13 9 million people in the US The presentation of ACS is varied and symptoms may develop suddenly with acute myocardial infarction AMI being the most dramatic of presentations Annually AMI affects approximately 1 1 million people in the US with a 30 mortality rate The World Health Organization recommends diagnosis of AMI with a positive indication on at least two of these three criteria 1 patient history and physical examination 2 electrocardiogram 3 changes in blood levels of cardiac protein markers Patient history and physical examination are critical and must be considered but often provide insufficient information to differentiate AMI from other cardiac abnormalities Electrocardiogram together with the patient history and physical examination is useful but diagnostic in only about 50 of AMI patients ST segment elevation or depression and Q wave formation are typical indicators of AMI Patients hospitalized with ACS have been found to be high risk ST segment elevation myocardial infarction STEMI or moderate high risk non ST segment elevation myocardial infarction NSTEMI patients However it may be difficult to diagnose AMI in NS
17. the whole blood The plasma from the spiked blood was 2 fold serially diluted into troponin negative plasma for a total of ten values spanning the measuring range of the StatusFirst Tnl test Each level was tested in ten replicates The data were analyzed in accordance with CLSI EP6 A and demonstrated linearity of the test device across the measuring range from 0 063 to 32 ng mL The data are shown below in Table 1 Table 1 Linearity of StatusFirst Tnl test Expected conc Measured conc Recovery ng mL ng mL 113 8 116 8 104 4 109 2 102 1 107 6 102 0 104 9 109 0 101 8 Page 8 Precision The precision of the StatusFirst Tnl test device was determined using samples where Tnl from NIST was spiked at four concentrations Table 2 The within day and total precision studies were performed in two runs per day in six replicates per run at each concentration level for 15 days with three DXpress Readers The within run and total variances and coefficients of variation CVs were computed according to CLSI guideline EP5 A Table 2 Precision Data Mean level Average Within run Average Total ng mL Std dev ng mL CV Std dev ng mL CV CROSS REACTIVITY AND INTERFERING SUBSTANCES Potentially Cross Reactive Substances The following cardiac proteins and peptides were tested for potential cross reactivity in the StatusFirst Tnl test device at the
18. ts with AMI The temporal relation of release of cTnl into the serum has been investigated and compared to the other established cardiac markers such as creatine kinase MB CK MB myoglobin and Troponin T Following AMI cTnl is released into the circulation with levels exceeding the upper reference limit of normal within 4 6 hours and peak levels are reached after 12 24 hours This early release profile is similar to CK MB However CK MB levels return to normal values in about 72 hours while levels of cTnl remain elevated for 5 7 days The release of cTnl into the blood involves myocardial damage primarily as a result of AMI however such damage may occur from causes other than AMI such as unstable angina congestive heart failure and ischaemic damage due to coronary artery by pass surgery Further the distinct structure of cTnl and the availability of highly specific detection methods for cTnl have increased the utility of this marker for the diagnosis of AMI in complex clinical conditions that involve skeletal muscle damage The high specificity of cTnl measurements for the identification of myocardial damage has been demonstrated in conditions such as the perioperative period after marathon runs and following blunt chest trauma Levels of other non cardiac specific protein markers such as CK MB and myoglobin are typically elevated in the blood following such skeletal muscle damage Principle The StatusFirst Tnl test
19. usFirst w EDTA WB StatusFirst w EDTA Plasma StatusFirst w Li Heparin WB StatusFirst w Li Heparin Plasma o D i gt 2 2 12 c o o i I I 0 4 0 6 1 Specificity Table 5 Area under Curve AUC with different sample types Sample Sample Numbers Tested 95 Conf Interval Types EDTA WB 94 90 EDTA Plasma 92 90 Li Heparin WB 89 94 Li Heparin Plasma 93 93 Page 12 To determine the best cutoff a series of presumed cutoff values were set up and the corresponding sensitivities and specificities were compared Table 6 Table 6 Sensitivity and Specificity Compared to Access AccuTnI StatusFirst EDTA WB EDTA Plasma Li Heparin WB Li Heparin Plasma i ae 100 a a a 99 94 86 81 A cutoff of 0 2 ng mL Tnl is recommended for diagnosis of AMI as this yields optimal performance of 99 sensitivity and gt 96 specificity relative to Access AccuTnI However laboratories should establish their own diagnostic cutoff concentration based on the clinical practice at their respective institutions Correlation of StatusFirst Tnl vs Access AccuTnI and Triage Cardiac Panel The following correlation equations were obtained by comparing StatusFirst Tnl to Access AccuTnI and Triage Cardiac Panel Since each company uses different TnI standard material for the test calibration the correlation factor would differ StatusFirst Tnl uses NIST standard
20. y difference is that RUN PATIENT requires a Patient ID whereas EXTERNAL QC requires a Sample ID Uploading Lot Specific Calibration Information using the Data Chip When you start to use a new lot of test devices the Reader will prompt the operator to insert the Data Chip that is supplied in the test kit Insert the Data Chip with the corresponding test device lot number select OK to continue and follow the prompts Page 5 Testing Patient Sample Procedure There are three modes of testing patient samples Read Now Batch Scheduler The operator is free to choose the mode most suitable for the particular task at hand The Scheduler mode is described below as an example 1 Open the pouch and remove the test device 2 Label the test device with the patient ID 3 Place the test device on a level surface 4 Testing the Patient Sample on the DXpress Reader e From the Main Menu select 1 RUN PATIENT e Scan or manually enter lot number barcode from the pouch e Confirm test device information type of test device and lot number as displayed on the screen and press ENTER e Scan or manually enter the Operator ID barcode e Scan or manually enter the Patient ID barcode e Select either Blood or Plasma Serum as appropriate e From the Incubation Time window select SCHEDULER e Within 30 seconds do the following Draw patient sample solution up to the line on the transfer pipette while holding the transfer pipette vertically and dis
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