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Preparation of HBV DNA reference standards and the experience of HBV NAT in Taiwan

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  1. Preparation of HBV DNA reference standards and the experience of HBV NAT in Taiwan Dr. Hwei-Fang Cheng Department of Health, Taiwan

  2. HBV DNA Reference Standards • National HBV DNA standard :106 IU/mL, 0.5 mL/vial (relative to NIBSC HBV NAT standard) • National HBV DNA working reagent :103 IU/mL, 1 mL/vial (relative to NIBSC HBV NAT working reagent) • Intended use for HBV NAT testing of blood and pooled plasma and biological research

  3. High titer (~109 IU/ml) HBV positive plasma(negative for anti-HIV,anti-HCV and NAT of HIV, HCV , HAV , and B19) a) genotype B b) virus inactivation: 60 oC for 1 hr c) dilute into ~106 IU/mL and ~103 IU/mL • Diluents: pooled human plasma (negative for HBsAg, anti-HIV, anti-HCV and NAT of HIV, HCV, HBV, HAV, and B19)

  4. Collaborative Study • Participating Labs (11 Labs) • US CBER/FDA • National Institute of Infectious Diseases (NIID) ,Japan • Canadian Blood Services • National Institute for Biological Standards and Control (NIBSC) • Victorian Infectious Diseases Reference Laboratory (VDRL), Australia • NTUH Hepatitis Research Center, Taiwan • Center of Disease Control, Taiwan • BFDA, Taiwan • Roche Diagnostics • Artus GmbH • Digene Diagnostic Technology

  5. 1st National HBV DNA Standard

  6. HBV DNA standard 10 9 8 7 6 Numbers 5 H2 4 H1 3 G K E2 2 D I1 B 1 E1 C J F A I2 0 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0 Estimated IU (log 10 /ml)

  7. +2SD Average 5.955 -2SD

  8. 1st National HBV Working Reagent *ND: not detectable

  9. Working reagent 10 9 8 7 Numbers 6 5 I1 4 H1 3 J F 2 H2 E2 I2 1 A C G D B 0 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0 Estimated IU (log10/ml)

  10. +2SD Average 2.953 -2SD

  11. Summary of National HBV DNA Standard and Working Reagent

  12. Stability Test – HBV DNA Standard at 5 ±3oC days

  13. Stability Test – HBV DNA Working Reagent at 5 ±3oC Cobas Amplicor days

  14. Summary • The assigned potency of HBV DNA standard and HBV working reagent are 106 IU/ml and 103 IU/ml, respectively. • Genotype of national HBV DNA standard and working reagents is genotype B; and the serotype is adw. • National HBV DNA standard and working reagent are stable at 4 ℃ for 4 weeks.

  15. HBV NAT Experience in Taiwan

  16. Blood center (Date collected) Tainan (March 2003) Taipei (June 2003) Taipei (June 2003) Units screened 1440 * 1296 * 516 ** H B V EIA-positive NAT-negative 0 0 0 EIA-negative NAT-positive 0 0 6 (1.2 %) EIA-positive NAT-positive 17 (1.2 %) 5 (0.4 %) 2 (0.4 %) (1)EIA:Murex HBsAg version 3.0 kit; Sensitivity: <0.2 ng (2)NAT *Pooled units test (12 units/pool); Roch High Pure Viral Nucleic Acid Kit (0.2 ml of extraction volume); In house nested PCR; Sensitivity of 22 IU/ml. ** Single unit test Ampliscreen HBV/Roch (1 ml of extraction volume) ; In house nested PCR; Sensitivity of 12 IU/ml