PROFICIENCY TESTING OF IN-HOUSE NAT ASSAYS USED FOR BLOOD SCREENING

XXI SoGAT International Working Group Meeting on the Standardization of NAT for the Safety Testing of Blood, Tissues and Organs

for Blood-Borne Pathogens

28 - 29 May 2009 Brussels, Belgium

Paul-Ehrlich-Institut, Langen, GermanyWHO Collaborating Centre for Quality Assurance of Blood Products

and in vitro Diagnostic Devices

Julia KreßMichael Chudy Micha Nübling

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German Regulations for NAT Blood Donor Screening

1999: first mandatory NAT was introduced for HCV (< 5.000 IU/mL ID) 2004: NAT was implemented for HIV-1 (< 10.000 IU/mL ID) HBV NAT is voluntarily performed by many blood donation services in-house developed NAT assays, CE-marked diagnostic assays (off-

label-use) and CE-marked NAT screening assays may be used assays are validated for the individual pool size (10 to 96 donations) validation studies are assessed by PEI NAT systems undergo regular external quality assessment programs

organised by PEI 2008: proficiency study for in-house NAT assays: HCV, HIV-1 and HBV

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In-house NAT Proficiency Study 2008: Objective

verification of the efficiency of in-house NAT assays for the detection of HCV, HIV-1 and HBV in blood donations regarding

analytical sensitivity genotype / subtype sensitivity specificity reproducibility

the participation in the proficiency study is mandatory for HCV and HIV-1 NATs, voluntary for HBV NAT

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In-house NAT Proficiency Study 2008: Study Design

detection limit of the HCV, HIV-1 and HBV NATs with respect to the pool size

testing panels: calibrated PEI reference preparation, two positive materials and negative plasma

HIV-1-samples: one missed by CTM v1, one with discrepant results 0.5 log dilution series starting with the required minimum sensitivity characterization of panels by CE-certified NAT screening systems:

cobas TaqScreen MPX Test, Procleix Ultrio Assay dilution of samples individually for each lab simulating the pool size encoding of labs and samples sample shipment on dry ice submission of qualitative results (reactive / non-reactive)

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In-house NAT Proficiency Study 2008: Participants

Invited laboratories: in-house NAT screening assays (non-CE-marked / CE-marked) CE-marked diagnostic assays used for screening (off-label-use) CE-marked screening system with large pool sizes (48, 96)

HCV: 30 labs16 in-house NATs, 16 diagnostic assays, 3 cobas TaqScreen MPX (5 labs: 2 different methods)

HIV-1: 30 labs15 in-house NATs, 15 diagnostic assays, 3 cobas TaqScreen MPX (3 labs: 2 different methods)

HBV: 21 labs14 in-house NATs, 6 diagnostic assays, 3 cobas TaqScreen MPX (2 labs: 2 different methods)

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In-house NAT Proficiency Study 2008: HCV Results1 2 3 4 5 6 7 8 9 10 11 12

PEI Ref #3443/04(genotype 1)

N2188(genotype 2)

N5022(genotype 4)

Neg. Plasma

Lab Test IU/mL 5.000 1.582 500 5.000 1.582 500 5.000 1.582 500Pool size 96

cobas TaqScreen MPX TestProcleix Ultrio Assay

2 cobas TaqScreen MPX Testcobas TaqScreen MPX TestCAP/COBAS TaqMan HCV Test

16 CAP/COBAS TaqMan HCV Test24 CAP/COBAS TaqMan HCV Test

in-house (1)in-house (2)

5 in-house6 in-house

17/1 in-house17/2 in-house

Pool size 4828 cobas TaqScreen MPX Test7 CAP/COBAS TaqMan HCV Test14 CAP/COBAS TaqMan HCV Test

HPS/COBAS TaqMan HCV Test 2.0COBAS AMPLICOR HCV Test 2.0

20 COBAS AMPLICOR HCV Test 2.013 in-house

in-house (1)in-house (2)

Pool size 4019 COBAS AMPLICOR HCV Test 2.0

Pool size 321 HPS/COBAS TaqMan HCV Test 2.0

Pool size 2415 COBAS AMPLICOR HCV Test 2.023 in-house11 in-house

Pool size 2022 COBAS AMPLICOR HCV Test 2.026 COBAS AMPLICOR HCV Test 2.031 COBAS AMPLICOR HCV Test 2.09 in-house

Pool size 1029 CAP/COBAS TaqMan HCV Test18 CAP/COBAS AMPLICOR HCV Test 2.0

in-house (1)in-house (2)

12 in-house21 in-house

reactive non-reactive

27

8

25

0 0

HCV

3

PEI

4

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In-house NAT Proficiency Study 2008: HIV-1 Results1 2 3 4 5 6 7 8 9 10 11 12

PEI Ref #3441/04(subtype B)

N4839(subtype B)

N4616(CRF02-AG/CRF04-cpx)

Neg. Plasma

Lab Test IU/mL 10.000 3.160 1.000 10.000 3.160 1.000 neat 1:3,16 1:10 0Pool size 96

cobas TaqScreen MPX TestProcleix Ultrio Assay

2 cobas TaqScreen MPX Testcobas TaqScreen MPX TestCAP/COBAS TaqMan HIV-1 Test

16 CAP/COBAS TaqMan HIV-1 Test24 CAP/COBAS TaqMan HIV-1 Test20 CA HIV-1 Monitor 1.5 UltraSens

in-house (1)in-house (2)

5 in-house6 in-house

17/1 in-house17/2 in-house

Pool size 4828 cobas TaqScreen MPX Test7 CAP/COBAS TaqMan HIV-1 Test14 CAP/COBAS TaqMan HIV-1 Test13 HPS/COBAS TaqMan HIV-1 Test27 HPS/COBAS TaqMan HIV-1 Test25 in-house

Pool size 4019 in-house

Pool size 321 HPS/COBAS TaqMan HIV-1 Test

Pool size 2415 CA HIV-1 Monitor v1.5 UltraSens22 Abbott RealTime HIV-1 Assay23 in-house11 in-house

Pool size 2026 CA HIV-1 Monitor v1.5 UltraSens31 CA HIV-1 Monitor v1.5 Standard9 in-house

Pool size 1018 CAP/COBAS TaqMan HIV-1 Test29 CAP/COBAS TaqMan HIV-1 Test

in-house (1)in-house (2)

12 in-house21 in-house

reactive non-reactive

0

8

HIV-1

3

PEI

4

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In-house NAT Proficiency Study 2008: HBV Results

1 2 3 5 6 7 8 9 10 11 12PEI Ref #3620/05

(genotype D)N4407

(genotype A)N4882

(genotype G)Neg. Plasma

Lab Test IU/mL 1.000 316 1.000 316 100 1.000 316 100 0Pool size 96

cobas TaqScreen MPX TestProcleix Ultrio Assay

2 cobas TaqScreen MPX Testcobas TaqScreen MPX TestCAP/COBAS TaqMan HBV Test

16 CAP/COBAS TaqMan HBV Test24 CAP/COBAS TaqMan HBV Test

in-house (1)in-house (2)

5 in-house6 in-house

17/1 in-house17/2 in-house

Pool size 4828 cobas TaqScreen MPX Test7 CAP/COBAS TaqMan HBV Test13 in-house

Pool size 4019 in-house

Poolsize 321 CAP/COBAS TaqMan HBV Test

Pool size 2423 in-house11 in-house

Pool size 209 in-house

Pool size 1029 CAP/COBAS TaqMan HBV Test12 in-house18 in-house21 in-house

reactive non-reactive

0

PEI

4

100

HBV

3

4

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In-house NAT Proficiency Study 2008: Conclusion

all participants meet the PEI sensitivity requirements

in most labs NAT assays show higher sensitivity than required

high specificity: only 1 false-positive result by 1 lab

HIV-1: CTM v1-missed sample was detected by other systems

due to high mutation rates there is a certain amount of risk that

infectious donations are missed by HIV-1 NATs

voluntary HBV NAT assays: high sensitivity and specificity

in-house NAT systems and off-label-use systems under proper

conditions are still suitable for blood donor screening