1. Linearity Panels HIV RNA, HCV RNA, HBV DNA, and CMV DNA
Mark Manak, Ph.D.
BBI Diagnostics, Inc.
A Division of SeraCare Life Sciences, Inc.
SoGAT XVIII
Bethesda, MD
May 24-25, 2005

2. Linearity Panel Design
Mimic clinical specimen
Whole virus in defibrinated plasma diluent
Challenges the entire analytical process
One negative, and Nine positive members
Series of sequential one log or half log dilutions
Results expressed as specified by kit manufacturers
Serial dilutions of the WHO International Standards analyzed simultaneously to assign IU values, where appropriate

3. Intended Use Assay Effectiveness Training
Evaluate sensitivity and dynamic range of quantitative
viral nucleic acid assays
Identifying consistency over a broad linear range
Establish calibrations vs. IU Standards
Verification of performance at kit lot changes
Training
Operator training program
Operator validation
Part of Laboratory Quality Assurance program

4. HCV RNA Linearity Panel (PHW804)
Diluent:
Normal human plasma
Negative for antibody to HIV-1/2, HBsAg, HCV, HTLV-I/II
Negative for HBsAg, HIV-1 RNA, HBV RNA and HCV DNA
Virus Spike:
High titer HCV RNA positive human plasma
HCV Genotype 1 by Innogenetics InnoLIPA Assay
Results on Data Sheet:
Roche COBAS Amplicor HCV Monitor v2.0
Bayer Versant HCV 3.0 bDNA Assay
Abbott LCx HCV Quantitative Assay
QuantaSure Plus HCV Roche Taqman ASR

7. HCV RNA Unit Conversions to IU
Roche COBAS: 2.2 copies/IU
Bayer Versant: 5.2 copies/IU
Abbott LCx: 4.3 copies/IU
Taqman: 0.6 copies/IU

8. HIV RNA Linearity Panel (PRD801)
Diluent:
Normal human plasma
Negative for anti-HIV-1/2, anti-HBsAg, anti-HCV, anti-HTLV-I/II
Negative for HBsAg, HIV-1 RNA, HBV RNA and HCV DNA
Virus Spike:
Culture supernatant: HIV-1 Type B 8E5 virus
Intact, but defective virus genome
Results on Data Sheet:
Roche COBAS Amplicor HIV Monitor v1.5
Roche COBAS Amplicor HIV Monitor v1.5 Ultra
Bayer Versant HIV-1 RNA bDNA 3.0 Assay
Chiron/Genprobe Procleix HIV-1 RNA Discriminatory Qualitative
BioMerieux Nuclisens HIV-1 QT

11. HBV DNA Linearity/Sensitivity Panel (PHD801)
Diluent:
Normal human plasma
Negative for antibody to HIV-1/2, HBsAg, HCV, HTLV-I/II
Negative for HBsAg, HIV-1 RNA, HBV RNA and HCV DNA
Virus Spike:
High titer HBV DNA positive human plasma
HBV DNA:Type A virus by Roche Taqman and Innogentics InnoLIPA
Serotype ad
Results on Data Sheet:
Roche COBAS Amplicor HBV
Digene HBV DNA Quantitative Assay
Bayer Versant HBV DNA Quantitative bDNA Assay
Taqman HBV DNA Quantitative Assay

13. HBV DNA Unit Conversions to IU
Roche COBAS: 1.2 copies/IU
Bayer Versant: 0.9 copies/IU
Taqman: 3.9 copies/IU
Digene: copies/IU
Digene: 1,200,000 IU/pg

14. CMV DNA Linearity/Sensitivity Panel (PTC801)
Diluent:
Normal human plasma
Negative for antibody to HIV-1/2, HBsAg, HCV, HTLV-I/II
Negative for HBsAg, HIV-1 RNA, HBV RNA and HCV DNA
Negative for anti-CMV and CMV DNA
Virus Spike:
Culture supernatant from CMV cultures
Results on Data Sheet:
Roche COBAS Amplicor CMV Monitor Test

16. Conclusions
BBI Linearity panels are made from natural virus in plasma diluent
Nine positive members spanning a broad range of virus concentrations and a negative member
Calibrated against WHO standards and expressed in IU/ml
Data from available commercial assays
Useful for monitoring linearity and dynamic range of assays
I would like to thank my co authors as well as my colleagues who assisted in this study.