FREND™ PSA Plus Prostate Specific Antigen
Contents
1. 8 65 9 24 818 8 67 7 74 8 555 0 526 6 1 8 0 106 9 3 0 375 13 30 12 33 12 71 13 91 12 61 11 58 12 740 0 802 63 12 8 99 9 4 0 500 18 34 16 89 18 22 16 07 17 56 18 57 17 608 0 972 5 5 17 0 103 6 5 0 625 21 44 22 49 19 39 19 32 20 67 22 78 21 015 1 490 7 1 21 3 98 9 6 0 750 23 78 25 42 27 13 26 88 24 91 25 93 25 675 1 255 4 9 25 5 100 7 7 0 875 28 04 34 33 27 12 29 51 27 36 26 38 28 790 2 913 10 1 29 8 96 8 High 1 000 39 98 35 22 31 74 27 43 28 32 40 09 33 797 5 561 16 5 34 0 99 4 lc Comparative Analysis A group of well characterized serum samples collected with IRB oversight from subjects with pathology verified prostate cancer and stored at 70 C under monitored conditions were analyzed for total PSA Results from the FREND PSA Plus on the FERND System y were compared to those obtained using the Tosoh ST AIA PACK PA assay x A total of 160 unique samples were analyzed in the study however only samples with tPSA results within the linearity of the FREND PSA Plus up to 25 0 ng mL n 143 were used in the comparative analysis Slope 0 9192 95 CI 0 8369 1 0014 Intercept 0 01179 95 CI 0 2763 0 2527 Correlation Coefficient R 0 9671 95 CI 0 9545 0 9763 Number of samples 143 Range of FREND PSA Plus values 0 04 29 99 ng mL Range of ST AIA PA values 0 00 25 86 ng mL 2 Precision 2a Precision Testing Single Lo2. FRENDTM System A minimum of at least two 2 levels of controls normal and abnormal should be used Individual laboratory policy will dictate exactly which control materials and lot numbers should be run the frequency with which controls are to be tested criteria for acceptance of the results and required corrective action to be taken if results do not meet laboratory criteria Do not assay patient samples on the FRENDTM System using the FRENDTM PSA Plus if quality control results do not give expected values Refer to your laboratory policies on how to determine acceptability of external control material results Quality Control Procedure External quality control materials to be assayed using the FRENDTM PSA Plus is defined by individual laboratory policy The assay procedure used to obtain results on the selected external quality control material is identical to that used when testing patient samples and follows below If lyophilized quality control material is to be used please reconstitute according to the manufacturer s instructions and allow the pellet to dissolve as stipulated until the material has gone into solution Be sure to mix gently but thoroughly before testing Refrigerated and or frozen control material should be treated as described below under preparation for specimen processing All materials should be at room temperature before use Specimen Processing e Preparation Remove from the refrigerator sufficient car
3. Geriatrics Society 2009 Vol 13 Mar pp 12 17 Chen Z Prestigiacomo A et al Purification and characterization of prostate specific antigen PSA complexed to alpha l antichymotrypsin potential reference material for international standardization of PSA immunoassays Clinical Chemistry 1995 41 9 1273 82 Wang M C et al 1981 Prostatic Antigen A New Potential Marker for Prostatic Cancer Prostate 2 89 Frankel A G et al 1982 Monoclonal Antibodies to a Human Prostate Antigen Cancer Res 42 3714 Papsidero L D et al 1980 A Prostate Antigen in Sera of Prostatic Cancer Patients Cancer Res 40 2428 Kuriyama M et al 1980 Quantitation of Prostate Specific Antigen in Serum by a Sensitive Enzyme Immunoassay Cancer Res 40 4658 Killian C S 1985 Prognostic Importance of Prostate Specific Antigen for Monitoring Patients with Stages B2 to D1 Prostate Cancer Cancer Res 45 886 Kuriyama M et al 1982 Multiple Marker Evaluation in Human Prostate Cancer with the Use of Tissue Specific Antigens J Nat Canc 68 99 Nahm MH and Goffman JW Heteroantibody Phantom of the Immunoassay Clinical Chemistry 1990 36 829 Boscato LM Stuart MC Heterophilic Antibodies A Problem for All Immunoassay Compared Clinical Chemistry 1988 33 1916 20 Young D 1990 Effects of Drugs on Clinical Laboratory Tests 3rd Edition Washington DC American Association for Clinical Chemistry Press Chan DW Bruzek DJ Oesterling JE et
4. reaction is read at various intervals during the analysis process blank readings are subtracted after which the net rate is automatically converted to total Prostate Specific Antigen concentration in ng mL based upon information stored on the PSA Code chip This result is then output on the screen and to the optional printer It is also stored in memory on the FREND System Screen Displays for Various Concentration Scenarios Displayed result Description By Date Time 2012 1 13 10 55 AM User ID Nano j Uer iD Nana PSA concentration Less than 0 10 ng mL lt 0 10 ng mL Date Time 2012 1 17 11 55 AM gt User D Nano PSA concentration Patient ID T10 Not less than 0 10 ng mL and not higher than 25 00 ng mL 9 73 ng mL Date Time 2012 1 13 11 30 AM User ID Nano PSA concentration Patient ID T 04 Higher than 25 00 ng mL gt 25 00 ng mL 12 Evaluation of Results Quality Control In order to monitor and evaluate the precision of the analytical performance it is recommended that commercially available control samples be assayed daily The minimum recommendations for the frequency of running internal control material are When beginning a new lot of cartridge three levels of controls are run in order to validate the calibration The three levels of controls are also repeated when certain service procedures are performed such as optical adjustment or change If any external quality control
5. the other FDA cleared PSA assay Based on the 95 confidence intervals there appears to be no differences between the concordances for the FREND PSA Plus assay and the other FDA cleared PSA assay Concordance FREND 95 CI FDA 95 CI PSA Plus cleared PSA Positive 77 8 69 8 to 85 3 81 5 73 85 to 88 7 Negative 64 1 55 1 to 71 5 64 1 55 1 to 71 5 Total 70 3 64 1 to 76 9 72 0 66 1 to 77 5 Confidence intervals are based on 10 000 resamples of the patient data 19 References ROTEN 11 12 13 14 15 16 Melissa Tanner Neil Kent Brian Smith Stephen Fletcher and Michelle Lewer Stability of common de Koning HJ Liem MK Baan CA Boer R Schr der FH Alexander FE Prostate cancer mortality reduction by screening power and time frame with complete enrollment in the European Randomized Screening for Prostate Cancer ERSPC trial International Journal Cancer 2002 98 268 73 Wan Ming Zhang Patrik Finne Jari Leinonen et al Characterization and immunological determination of the complex between prostate specific antigen and a2 macroglobulin Clinical Chemistry 1998 44 12 2471 2479 David L Woodrum et al Analytical performance of the Tandem R free PSA immunoassay measuring free prostate specific antigen Clinical Chemistry 1997 43 7 1203 1208 Yun Sik Kwak M D Appropriate use of Prostate Specific Antigen in Diagnosing Carcinoma of the Prostate Journal Korean
6. 15 of the expected is considered acceptable performance 1 2 Endogenous Interference These interference studies on endogenous substances were performed using the FREND PSA Plus on the FREND System according to the recommendations in the CLSI protocol EP7 A Added hemoglobin up to 500 mg dL does not interfere with the assay Average recovery when added to serum containing tPSA at 10 and 4 0 ng mL was 97 25 Added conjugated bilirubin up to 20 mg dL does not interfere with the assay Average recovery when added to serum contaiing tPSA at 1 0 and 4 0 ng mL was 98 2 Added gamma globulin Total Protein up to 5 0 g dL does not interfere with the assay Average recovery when added to serum containing tPSA at 1 0 and 4 0 ng mL was 106 3 Added triglyceride up to 3 g dL does not interfere with this assay Average recovery when added to serum containing tPSA at 1 0 and 4 0 ng mL was 101 5 Pharmaceutical Interference The following chart shows the interference studies performed using the FREND PSA Plus on the FREND system for various drugs that might be found in the serum plasma of men diagnosed with prostate cancer The concentrations of the drugs that were added to the test samples and controls at base concentration of tPSA of 1 0 ng mL and 4 0 ng mL are shown as are the recoveries on the far right side of the chart It is unlikely since the FREND PSA Plus method uses monoclonal antibodies that any substances without a te
7. NESPI FRPS O0I1E V 0 1 FREND PSA Plus Prostate Specific Antigen 1 Intended Use The FREND PSA Plus is designed for in vitro DIAGNOSTIC USE ONLY for the quantitative measurement of total Prostate Specific Antigen PSA in human serum heparinized plasma and EDTA plasma using the FREND System This device is indicated for the serial measurement of total PSA in serum heparinized plasma and EDTA plasma to be used as an aid in the management of patients with prostate cancer 2 Summary and Explanation of Test Prostate specific antigen PSA is a single chain glycoprotein with molecular weight of 34 kilodaltons 3 As a serine protease with chymotrypsin like activity PSA belongs to the kallikrein family In blood PSA exists as a free or complex form with protease inhibitors such as a I antichymotrypsin ACT Total PSA represents the sum of both free and complex forms PSA is uniquely associated with prostate tissues from normal inflamed or cancerous stages Elevated PSA in serum or plasma is found in patients with prostate cancer benign prostatic hypertrophy or inflammatory tissues Studies on a variety of PSA methods have shown that PSA can be useful as an indicator for the diagnosis and management of prostate cancer PSA has been found in normal benign hyperplastic and malignant prostatic tissue in metastatic prostatic carcinoma and also in prostatic fluid as well as in seminal fluid PSA is not found in any other tissue in
8. al Human Anti Murine Immunoassays Clinical Chemistry 1997 1916 20 Chemistry Analyte Stability Chart Barnes Jewish Hospital Apr 1999 Revised on Jul 2011 biochemical analytes in serum gel tubes subjected to various storage temperatures and times pre centrifugation 15 Annals of Clinical Biochemistry 2008 45 375 379 18 G Rajani Kumari and T Malati Stability of total and free prostate specific antigen in serum samples at different storage conditions Indian Journal of Clinical Biochemistry 2004 19 2 10 13 20 Glossary of Symbols Definition Do not reuse Use by YYYY MM DD Symbol Lot number Catalogue number N Warning or Caution Manufactured by ec rer Authorized representative in the Europe Community vD In vitro diagnostic medical device X Temperature limitation Contains sufficient for lt n gt tests NanoEnTek Inc Email www henryschein com requestinfo Website www henryschein com frend Mh tanufactured by NanoEntTek Inc H Q 12F 5 Digital ro 26 gil Guro gu Seoul 152 740 Korea Tel 82 2 6220 7940 Fax 82 2 6220 7721 Manufacturing site 851 14 Seohar ro Paltan myeon Hwaseong si Gyeonggi do 445 917 Korea US Branch NanoEnTek USA Inc 5627 Stoneridge Drive Suite 304 Pleasanton CA 94588 USA Tel 1 925 225 0108 1 888 988 0108 toll free Fax 1 925 225 0109 China SK Medical Beijing Co Ltd 26F SK Tower N
9. ase anti PSA antibodies PSA fluorescent particle immune complexes in the specimen are grabbed by the capture antibodies to form sandwich immune complexes The residual PSA unbound fluorescent nano particles conjugated with PSA antibodies pass through the test zone and bind to PSA antigens in the reference zone As the samples moves forward to the waste reservoir non specific binding components are washed away The intensity of fluorescence measured by a light source laser is proportional to the amount of total PSA in the original sample The result is calculated using information stored on the lot specific FREND PSA Plus Code chip and then is displayed on the FREND System screen A hard copy printout can be obtained if desired A ratio calculated between the Reference zone and the Test zone corrects for test to test variations Total PSA concentration in a sample analyzed with the FRENDTM PSA Plus on the FRENDTM System correlates directly with the fluorescence intensity the higher the tPSA concentration the greater the fluorescence The FREND PSA Plus has a measuring range determined as 0 1ng mL to 25 0ng mL The FRENDTM PSA Plus uses single use transparent plastic cartridges in which all required reagents are stored within the cartridge itself All that is added by the user is a 30uL test sample The cartridge is inserted into the FREND System in a prescribed fashion indicated with a black arrow on the cartridge The reaction is read mu
10. d serum samples or samples containing particulate matter such as fibrin clots or strands should be centrifuged before being tested Prior to assay slowly bring frozen samples to room temperature 18 25 C and mix gently but thoroughly before testing 9 Procedure 1 2 Reagent Preparation Cartridges There is no reagent preparation required to measure tPSA using the FREND PSA Plus cartridge on the FREND System However the cartridges needed for a particular run should be removed from the refrigerator and allowed to reach room temperature for 15 30 minutes before they are used Calibration The calibrators used during the cartridge manufacture process to create the information placed electronically on the FREND PSA Plus Code chip are prepared gravimetrically and are compared to international reference standards WHO International Prostate Specific Antigen 90 10 NIBSC code 96 670 However for the end user there is no need for calibration as is generally performed on other automated laboratory equipment All calibration statistics and information have been electronically stored on the FREND PSA Plus Code chip included in each box of FREND PSA Plus cartridge The FREND PSA Plus Code chip is specific for that manufactured lot of FREND PSA Plus T The appropriateness of the calibration information should always be checked by running sufficient external quality control materials as samples to verify that the res
11. e assay except in the case of gross lipemia where interference with the lateral flow of the sample in the cartridge may occur Specimens from patients who have received preparations of mouse monoclonal antibodies for diagnosis or therapy may contain human anti mouse antibodies HAMA Such specimens may show falsely elevated or decreased PSA values Certain medications may interfere with assay performance All results should be interpreted with respect to the clinical picture of the patient The concentration of tPSA in a given sample determined with assays from different manufacturers can vary due to differences in assay methods calibration and reagent specificity Please refer to the Specimen Collection and Handling Warnings and Precautions Storage and Stability and Procedural Notes sections in this insert sheet Clinical results must be interpreted with regard to medications administered to the patient The ability of the assay to detect both free and complexed forms of total PSA free PSA complexed with alpha 1 antichymotrypsin on an equal molar basis equimolarity has not been established 14 Expected Values As with every clinical diagnostic test a reference interval corresponding to the characteristics of the population being tested should be determined by each laboratory The FREND PSA Plus on the FREND System is to be used on serial blood samples to manage patients with prostate cancer Testing of ambulatory male
12. haracteristics Performance characteristics were evaluated for the FREND PSA Plus as follows 1 Accuracy la Recovery Known samples of PSA were added to a female serum specimen 0 01 ng mL The concentration of PSA was determined before and after the addition of the exogenous PSA and the percent recovery was calculated Spiked Recovery FREND PSA Plus Concentration added Observed concentration Recovery ng mL ng mL 1 08 1 06 98 3 1 09 100 7 1 04 96 7 4 34 4 42 101 8 4 35 100 3 4 27 98 4 12 81 13 58 106 0 12 04 94 0 11 82 92 3 25 53 24 26 95 0 24 67 96 6 26 90 105 4 1b Dilution Linearity Specimens from a high concentration pool 34 ng mL tPSA were diluted with a low concentration pool following instructions in the CLSI EP6 A document Correlation with the expected linearity showed R 0 9992 Dilution Linearity FRENDTM PSA Plus 10 w oO r oO 3 E uw x a 4 a P A a w N oO y 0 9866x 0 1324 R 0 9992 20 30 Expected value ng ml Dilution Linearity Data FREND PSA Plus No Dilution TEST TEST TEST TEST TEST TEST MEAN SD CV Expected 1 2 3 4 5 6 ng mL Value Recovery Blank 0 000 0 00 0 00 0 10 0 00 0 00 0 10 0 033 0 0 1 0 125 4 63 4 65 4 08 4 27 4 59 4 26 4 413 0 241 5 5 4 3 103 8 2 0 250 8 85
13. impression was then compared to significant changes in the PSA concentration of each sample as measured by both the test device FREND PSA Plus and the predicate device FDA Cleared Assay The results of these comparisons are shown below where clinical status was divided into two groups those visit pairs showing progression and those showing no disease progression per the visit to visit status 1 Samples with no Progression Negative Concordance Other FDA Cleared PSA Assay FREND PSA Plus gt 8 5 lt 8 5 Total 20 31 15 46 lt 20 15 67 82 Total 46 82 128 NC FREND PSA Plus 0 641 NC AIA PACK PSA 0 641 Different 0 000 There is no difference in the Negative Concordance NC between the two assays 2 Samples with Progression Positive Concordance Other FDA Cleared PSA Assay FREND PSA Plus gt 8 5 lt 8 5 Total 20 83 1 84 lt 20 5 19 24 Total 88 20 108 14 NC FRENDTM PSA Plus 0 778 NC AIA PACK PSA 0 815 Different 0 037 A McNemar test of the paired data in the above table yields a p value of 0 218 The associated 95 confidence interval for the true difference is 0 0157 to 0 0551 Both estimators agree there are no statistically significant differences in the Positive Concordances PC between the two assays Below is chart comparing the concordances of the FRENDTM PSA Plus assay and
14. is clearly indicated on the product box and the cartridges Materials Storage condition Catalogue number FREND PSA Plus cartridges Refrigerator temperature 2 8 C FRPS 025 Pipette tips Room temperature 18 25 C None 8 Specimen Collection and Handling Serum or plasma heparinized or EDTA only is required for the assay Citrated plasma SHOULD NOT BE USED No special patient preparation is necessary To use serum a blood sample is collected aseptically without additives by venous puncture After allowing the sample to clot for 30 minutes at room temperature the collection tube should be centrifuged for 10 minutes at 3 000 rpm For heparinized or EDTA plasma a venous blood sample is collected aseptically with the designated additive The plasma should be separated from the packed cells as soon as possible Prostatic manipulation has been shown to affect the PSA results so samples should be drawn before any prostatic procedures such as DRE prostatic massage and TRUS are performed Samples may be stored at 2 8 C for up to 6 hours prior to analysis If the analysis is scheduled to be done at some later time the sample should be stored frozen at 20 C or below for future use Sample stability study was performed at 20 C only for three weeks by manufacturer but there are many literature available showing that total PSA kept frozen is stable for 3 months or longer 17 8 Repeated freeze thaw cycles should be avoided Turbi
15. ltiple times as the sample moves via capillary action through the cartridge This type of assay system is sometimes referred to as one which incorporates laminar flow 4 Material Provided FREND PSA Plus Catalogue number 25 FREND PSA Plus cartridges FRPS 025 30 Disposable pipette tips micro pipettor provided 01 FRENDTM PSA Plus Code chip 01 FRENDTM PSA Plus Package Insert One Cartridge contains Monoclonal anti PSA1 48 9 6 ng Monoclonal anti PSA2 144 28 8 ng Fluorescent particle 2 4 0 48 ug 5 Materials Required But Not Provided The following materials are not provided with the cartridge but are required to perform Prostate Specific Antigen analysis using the FREND PSA Plus on the FREND System They are available separately from NanoEnTek Instrument Catalogue number FREND System F10 6 Warnings and Precautions The FREND PSA Plus cartridges are intended for in vitro diagnostic use only PSA Plus cartridges are only to be used on the FREND System Allow cartridges to come to room temperature for 15 30 minutes prior to use Avoid cross contamination between samples by using a new pipette tip for each new specimen Avoid high humidity direct sunlight or heat in the area used for cartridge storage Inaccurate results are possible if the sample used is contaminated in any way Using specimens containing clotted fibrin could result in erroneous results Over or under loading the cartridge with sample may
16. matically be expelled and the results displayed Caution Do not remove power from the FREND System while a cartridge is in the reading chamber This may cause a system error 10 If the FREND System is connected to the optional printer press the Print button and the results will be output on the printer paper 11 For more detailed instructions please refer to the FREND System User Manual 10 Procedural Notes If a specimen Prostate Specific Antigen concentration is found to be greater than the linearity limit of the assay of 25 0 ng mL and a definitive result is required the specimen should be diluted with female sera that has been previously measured on the FREND PSA Plus and found to contain lt 0 1 ng mL tPSA and then re assayed according to the Assay Procedure The recommended dilution for samples with an initial result of gt 25 0 ng mL is 1 10 or 1 50 It is desirable to dilute the sample so that the diluted sample reads between 2 and 20 ng mL Dilutions must be made manually and the final result on the diluted sample calculated manually by multiplying the result obtained on the diluted sample by the dilution factor 11 Calculation of Results The FREND System performs all sample and cartridge handling operations automatically within the cartridge once the sample has been manually added to the sample well in the cartridge and the cartridge placed into the FREND System The rate of fluorescence produced by the
17. men and it is not produced by cancers originating in the lung colon rectum stomach pancreas or thyroid Though increased concentrations of PSA are found in the serum of patients with benign prostate hyperplasia BPH prostatitis and prostate infections and inflammation they are also found in patients with cancer of the prostate PSA measurement is an essential tool in assessing the status of disease in patients with prostate cancer when serial samples are measured over time The clinical value realized by monitoring tPSA concentrations in patients with prostate cancer regardless of the treatment regimen is well known Since the mid 1980 s there has been a growing body of literature concerning the utility of Prostate Specific Antigen PSA for both the monitoring and detection of prostate cancer CaP 3 Principle of the Assay The FREND PSA Plus is a rapid quantitative sandwich immunoassay using fluorescent nanoparticles which measures the concentration of total PSA Thirty uL of patient serum or plasma heparin or EDTA only is manually presented to the inlet on the individual single unit test cartridge where it is mixed with fluorescent nano particles conjugated with PSA antibodies PSA molecules in the specimen bind to conjugated antibodies to form immune complexes which then move by capillary action through the reagent cartridge channel to the detection area When the specimen reaches the test zone it hydrates dried solid ph
18. o 6 Jia Jianguomenwai Avenue Chaoyang district Beijing 100022 P R China Tel 86 10 5920 7844 Fax 86 10 5920 5697 16 mc MT Promedt Consulting GmbH Altenhofstrasse 80 66386 St Ingbert Germany C Erz 17
19. result in inaccurate results Cartridges should not be frozen Human specimens are not used in the preparation of this product however since human specimens will be used for samples and other quality control products in the lab may be derived from human materials please use standard laboratory safety procedures when handling all specimens and controls Do not use the cartridges beyond the expiration date on the pouch Do not use the cartridge ifthe pouch is damaged or the seal is broken Perform testing as specified in the Package Insert and User Manual PSA Plus cartridges are disposable single use devices Do not reuse them under any circumstances Keep the cartridge sealed in the pouch until just ready for use Use the cartridge immediately after opening its pouch Wear disposable gloves when handling the cartridges and the samples Wash hands thoroughly and often handling reagent cartridges or samples PSA Plus has been designed so that the high dose hook effect is not a problem for the vast majority of samples Samples with PSA concentration between 25 and 1 200 ng mL will read gt 25 ng mL The hook effect phenomenon may occur only at PSA concentration gt 1 200 ng mL 7 Storage and Stability All unopened materials are stable until the expiration date on the label when stored at the specified temperature Cartridge stability has been demonstrated for twelve months from the date of manufacture The expiration date
20. rtiary structure similar to PSA would interfere The testing showed that indeed there was no significant interference from the tested drugs that would affect the interpretation of a tPSA result as assayed on the FREND PSA Plus Interference Study Results for FREND PSA Plus on the FREND System No Substrate Concentration Average Recovery 1 Flutamide 10 pg mL 94 50 2 Diethylstilbestrol DES 5 pg mL 103 80 3 Goserelin 40 ng mL 103 20 4 Tamsulosin 100 ng mL 98 85 5 Acetaminophen 250 ng mL 100 45 6 Acetylsalicylic acid 600 pg mL 95 85 7 Leuprolide 275 ng mL 101 50 8 Ibuprofen 500 ug mL 102 25 9 Finasteride 250 ng mL 93 60 10 Docetaxel 10 pg mL 114 45 18 Serial Measurements and concordance with medical Status Since the FREND PSA Plus indication for use is that the assay results will be used as a tool in managing the care for patients with prostate cancer it is imperative that the changes in the marker are compared to clinical status changes to determine the efficacy of the test Therefore as an important part of the clinical studies performed to 13 characterize the FRENDTM PSA Plus serial samples collected longitudinally from patients previously diagnosed with prostate cancer and treated in a variety of ways over the clinical course of their disease including prostatectomy radioactive seeds external beam radiation chemotherapy hormone therapy alone or in combination were a
21. s of this study are shown below Category Other FDA cleared PSA PSA Plus on FREND System Number of samples n 196 196 Mean x 0 71 ng mL 0 83 ng mL SD 0 38 ng mL 0 43 ng mL Range of values 0 00 2 03 ng mL 0 02 2 69 ng mL Median 0 66 ng mL 0 77 ng mL Expected Values for Management of Patients with Prostate Cancer Distribution of Serum FREND PSA Plus concentrations Healthy Benign and Various Malignant Disease States N 0 4 0 4 1 10 0 10 1 20 0 20 1 40 0 gt 40 0 ng mL ng mL ng mL ng mL ng mL Healthy Subjects 196 Men gt 50 yrs 196 100 0 0 0 0 Benign Disease Cond 410 Benign Prostate 104 56 73 25 96 11 54 3 85 1 92 Diabetes 97 95 88 3 09 1 03 0 00 0 00 HTN Heart Disease 102 95 10 4 90 0 00 0 00 0 00 Benign GI 107 94 4 4 67 0 00 0 93 0 00 Malignant Diseases 302 Prostate Cancer 85 40 00 38 82 12 95 2 35 5 88 Gleason Score 5 6 43 51 16 44 19 2 38 2 38 0 Gleason Score 7 31 35 48 38 72 19 35 0 6 45 Gleason Score 8 9 11 9 09 18 18 36 36 9 09 27 21 Lung Liver Cancer 52 98 08 0 1 92 0 0 GB Gastric Pancreatic 31 100 0 0 0 0 Colorectal Cancer 89 94 38 4 49 1 13 0 0 Other Cancers 45 97 78 2 22 0 0 0 Total Subjects 908 Treated and untreated subjects Serial samples are not included in this cohort 16 Performance C
22. sample values are out of the acceptable range it will be necessary to investigate the problem before reporting patient results to assure there is not an instrument or software malfunction Each laboratory operates under a different set of regulations Every laboratory must follow the standardized procedures acceptable to the regulatory agencies to whom the laboratory is responsible 13 Limitations of the Procedure When used for diagnostic purposes the results obtained from this assay should be used in conjunction with other data e g symptoms results of other tests clinical impressions medical history therapy etc The FREND System paired with a FREND PSA Plus cartridge is programmed to report 25 0 ng mL as the highest concentration of PSA measurable without dilution The lowest measurable concentration is 0 1 ng mL the assay sensitivity limit Heterophilic antibodies in a sample have the potential to cause interference in immunoassay system Infrequently PSA level may appear elevated due to heterophilic antibodies present in the patient s serum or plasma or to nonspecific protein binding If the PSA level is inconsistent with clinical evidence additional PSA testing is suggested to confirm the results Although hemolysis has an insignificant effect on the assay hemolyzed samples may indicate mistreatment of a specimen prior to assay and results should be interpreted with caution Lipemia has an insignificant effect on th
23. ssayed for tPSA with the FRENDTM PSA Plus on the FREND System The same samples were also measured for tPSA by another FDA cleared method For each point to point in as sample serial set the change in the tPSA concentration was compared to the change in the clinical status of the patients as measured by other laboratory tests patient interviews physical examinations and imaging studies of a variety of types and recorded on a Clinical Report Form These changes in the tPSA marker concentration were defined as significant or not by multiplying the overall CV of the assay at the midrange as determined by the test imprecision study by a factor of 2 5 to define a percentage change different from what would be expected because of assay imprecision For the FRENDTM PSA Plus assay with an overall mid range CV of 8 5 significance was set at a change in excess of 20 Any increase in value from one time period to the next that did not exceed 20 was logged as lt 20 change For the other FDA cleared method significance was set at a change gt 8 5 This was calculated using that method s published overall mid range CV of 3 4 x 2 5 Physician s impressions regarding subject s disease status at each blood draw were recorded on CRFs for all serial PSA samples The status information at one visit was compared with the status at the next and a change in status was determined for the visit pair The change in clinical status from the physician s
24. subjects fifty years old and older who reported themselves as healthy without any known illnesses diseases or conditions was performed using both the FREND PSA Plus on the FREND System and another commercially available PSA method The currently accepted reference interval for tPSA of up to 4 0 ng mL was validated for both systems As is true for all PSA methods no tPSA results can be interpreted as being definitive for the presence or absence of prostate cancer Patients with levels of PSA within the reference interval found in apparently healthy subjects may have prostate cancer patients with levels exceeding those in the reference interval may be prostate cancer free Results from the FRENDTM PSA Plus on the FRENDTM System should be interpreted in the light of other clinical findings and diagnostic procedures such as DRE various imaging studies etc since certain treatments can cause PSA values to decrease by virtue of the treatment while the cancer is still progressing 15 Reference Ranges The interval given here was determined in serum samples from 196 apparently healthy male subjects from the age of 50 71 years Category Men Number of samples n 196 Reference interval 0 4 0 ng mL In this study greater than 99 of the healthy subjects had serum PSA concentrations less than or equal to 4 0 ng mL by FREND PSA Plus on both the FREND System and another commercially available tPSA fluorescent assay Result
25. t and Single Site Precision was determined as described in the CLSI protocol EP5 A Three clinical samples across the measuring range were assayed in replicates of two at two separate times per day for twenty days using a single lot of FRENDTM PSA Plus cartridge The findings follow showing repeatability between run between day and within laboratory precision data Sample Mean PSA Repeatability Between run Between day Within ng mL laboratory SD CV SD CV SD CV SD CV 1 0 098 0 013 12 8 0 005 5 5 0 004 3 7 0 014 14 4 2 4 321 0 248 5 7 0 054 1 2 0 089 2 1 0 269 6 2 3 12 735 0 636 5 0 0 405 3 2 0 102 0 8 0 761 6 0 4 25 462 1 278 5 0 0 668 2 6 0 321 1 3 1 477 5 8 2b Precision Testing Multiple Lots and Multiple Sites Three different lots of FREND PSA Plus were evaluated at three geographically diverse sites Four replicates each of Material A Material B and Material C and two replicates of QC 1 QC 2 and QC 3 were evaluated in each of two runs performed for five days at each site A total of 40 results on each material were generated at each of the three sites yielding a grand total of 120 replicates of each material The data was analyzed using a CLSI format from EP5 A2 for an ANOVA analysis Instrument to Instrument is the same as Site to Site in this scenario As can be seen from the table below the largest source of variation is the cartridge which
26. tridge of FREND PSA Plus to test the number of patient samples and required external quality control materials Allow the cartridges to come to room temperature for 15 30 minutes prior to the start of the testing sequence If using refrigerated patient samples remove those from the refrigerator and allow to them to come to room temperature prior to testing If frozen samples will be utilized be sure these are removed from the freezer thawed naturally and then mixed gently but thoroughly prior to testing There are no other reagents or sample preparations necessary e Assay Procedure 1 Prepare the FREND PSA Plus and specimen 2 Record the Sample ID on the cartridge in the designated area 3 Drop the sample 30uL into the sample inlet on the cartridge using the FREND System pipettor with a fresh pipette tip 4 Press the Test button on the Main screen of the FREND System 5 The screen of FREND System moves to the Patient ID screen automatically 6 Type the Patient ID and press the Enter button to begin the test 7 Insert the cartridge into the cartridge slot using the cartridge arrows as a guide Caution Please check the direction of the cartridge before insertion and assure the insertion is complete 8 When the reaction in the cartridges is complete in 6 minutes the FREND System will automatically begin the reading process 9 When the measurements are completed the cartridge will auto
27. ults obtained for tPSA on the FREND System using the FREND PSA Plus cartridges of a particular lot met the laboratory criterion for acceptability e PSA Plus Code Chip Installation Please refer to the FREND System User Manual for more detailed instructions relative to the Code chip installation Abbreviated instructions follow here 1 Insert the FREND System electrical cord into an appropriate outlet 2 Insert the Code chip into the code chip slot at the rear of the FREND System following the arrows 3 Press the Setup button on the Main screen 4 Press the Code chip button on the Setup screen 5 The information embedded on the FREND PSA Plus Code chip is automatically saved on the FREND System 6 When the Code chip installation is completed press the OK button to go to the Setup screen 7 Press the Item button on the Setup screen 8 Check the FRND PSA Plus cartridge lot number and the installation date of the Code chip 9 Press the Home button to go to the Main screen to begin running external quality control and patient samples 3 4 Quality Control e Commercially Available Controls C Commercially available controls from a variety of manufacturers are available that contain tPSA as a measured analyte It is recommended that these external controls be run at least once per day when testing is scheduled for FRENDTM PSA Plus on the
28. would be the expected result The FREND PSA Plus cartridge is a single use cartridge that contains all the reagents within the cartridge necessary to support the reactions CV by Material Material Variation MATA MAT B MAT C Qc 1 QC 2 QC 3 Source 0 29 ng mL 3 67 ng mL 18 33 ng mL 0 30 ng mL 2 93 ng mL 20 25 ng mL Site to Site 3 50 1 57 1 67 3 47 1 61 2 06 Day to Day 0 00 0 99 1 21 0 00 0 00 0 00 Lot to Lot 9 12 3 16 7 01 6 08 4 30 6 00 Inter cartridge 18 45 681 7 94 20 03 6 17 7 49 Total 20 87 7 14 10 79 21 22 7 69 9 81 3 Specificity The following substances were evaluated for potential cross reactivity with the FREND PSA Plus at the concentrations indicated below Testing was done according to the instructions recommended by CLSI protocol EP7 A No significant cross reactivity was found Specificity of FREND PSA Plus No Substrate Concentration 1 PAP 10 0 ng mL Prostatic Acid Phosphatase 2 Kallikrein 15 0 ng mL 4 Analytical Sensitivity The Limit of Detection LoD for the FREND PSA Plus was determined using the CLSI EP17 A protocol The analytical sensitivity of the FREND PSA Plus was determined to be 0 1 ng mL 17 Interference 12 Interference is defined for purposes of this study to be recovery outside of 15 of the known specimen mean concentration In other words recovery from 85 to 1
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