SoGAT XVII/Paris, 2004

Proficiency TestingProficiency Testingof In-House Developed HIV-1 NAT of In-House Developed HIV-1 NAT

for Blood Screeningfor Blood Screening

Michael Chudy, Paul-Ehrlich-InstitutDivision of Virology, Section of Molecular Pathology

Langen, Germany; Email: chumi@pei.de

PEI

German Regulations for Blood Components German Regulations for Blood Components and FFP: Implementation of HIV-1 NATand FFP: Implementation of HIV-1 NAT

BAnz. [Fed. Gazette] No 90, 9506; 05/16/2001 detection limit of HIV-1 RNA: 10,000 IU/ml for

single donation prospective implementation 01/01/2002

08/27/2001: PEI announcement HIV-1 subtypes: M group PEI collaborative study of in-house NAT assays

for detection of HIV-1 and HCV RNA;participation on voluntary basis (10/01/2001)

HIV-1 NAT implementation postponed

PEI

HCV Samples HIV-1 Samples

1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

2 3 4 5 A E C D

5.000 5.000 1.580 15.800 5.000 5.000 5.000 5.000 5.000 5.000 31.600 10.000 10.000 3.160 10.000 10.000 10.000 10.000 10.000 10.000

1 96

3 96

18 96

7 96

9 96

12 96

20 96

6 48

14 1(48)

8 48

17 48

2 20

19 20

24 20

21 12

13 10

15 10

16 10

23 10

4 1

5 1

10 1

22 1

pos neg

NegWHO PEI

NegLab CodePool Size

WHO PEI

Collaborative Study: In-house NAT AssaysCollaborative Study: In-house NAT Assays

PEI

Collaborative Study 2001: Collaborative Study 2001: ConclusionsConclusions

HCV NAT (in-house): homogeneous results (sensitivity, specificity, and

proficiency) no differences between SD- and MP-testing

HIV-1 NAT (in-house): results less homogeneous further improvements and validation regular proficiency studies for in-house NAT

assays organised by PEI acceptance: successful participation

PEI

German Regulations for Blood Components German Regulations for Blood Components and FFP: Implementation of HIV-1 NATand FFP: Implementation of HIV-1 NAT

BAnz. [Fed. Gazette] No 103, 12269; 06/05/2003 implementation 05/01/2004 validation of in-house assays acceptance of in-house assays: successful

participation in PEI proficiency study

PEI

Proficiency Study for In-house HIV-1 Proficiency Study for In-house HIV-1 NAT Assays: : ObjectiveNAT Assays: : Objective

Evaluation of in-house NAT assays for detection of HIV-1 in blood donations:

analytical sensitivity subtype sensitivity specificity proficiency

PEI

Proficiency Study for In-house HIV-1 Proficiency Study for In-house HIV-1 NAT Assays: Study DesignNAT Assays: Study Design

• Focus on the detection limit:• 10,000 IU HIV1-RNA/ml single donation

• Materials• Reference preparations (WHO: 31,600, 10,000

2x, 3,160 IU/ml/PEI: 10,000 IU/ml)• HIV-1 subtypes (A, C, CRF_AE, G, H; each

10,000 IU/ml)• NHP pool (2x)

• Simulation of the pool size (volume, dilution)• Lab and sample encoding• Samples frozen at -80°C/shipment on dry ice• Results: reactive/non-reactive (data sheet)

PEI

Proficiency Study for In-house HIV-1 Proficiency Study for In-house HIV-1 NAT Assays: ParticipantsNAT Assays: Participants

Overall 30 participants 29 labs: blood screening; 2 labs only pooling/1

lab: manufacturer of plasma prod. SD 1x; MP-8 1x; MP-10 3x; MP-15 2x, MP-20

4x, MP-24 2x, MP-40 1x, MP-48 5x; and MP-96 11 x

7 labs with virus enrichment NA extraction: 14 labs Qiagen/5 labs Roche/4

labs NucliSens/5 labs other 23 real-time PCR (20x TaqMan/3x LC Hybr.) 2x CA HIV-1 Monitor/3x RT-PCR +Gene

ScanningPEI

HIV-1 Samples

1 2 3 4 5 6 7 8 9 10 11 12PEI * A C E G H

31.600 10.000 10.000 3.160 10.000 10.000 10.000 10.000 10.000 10.000

2 963 96

6 969 96

13 9615 96

21 9629 96 (1)

30 96 (1)32 96

11 48

14 4820 48

23 4827 48

12 40

4 24

31 24

1 205 20

25 2026 20

17 15

18 15

8 1016 10

22 10

7 8

28 1

pos neg

19 96

WHONeg

Lab CodePool Size

Proficiency Study In-house HIV-1 NAT: ResultsProficiency Study In-house HIV-1 NAT: Results

Proficiency Study for In-house HIV-1 Proficiency Study for In-house HIV-1 NAT Assays (2003): ConclusionsNAT Assays (2003): Conclusions

PEI

• Results more homogeneous compared to the results of the study in 2001

• No differences in respect with the used pool size

• Due to the test results, 2 test labs switched to commercial assays

• Assessment of in-house assays: validation studies and successful participation in proficiency study

• Conclusion: In-house assays are suitable for blood screening

• CE-marking of in-house assays: open question