1. SoGAT 2005 Donor screening for parvovirus B19 antibodies: Reducing or eliminating the risk of transmission SoGAT 2005 Gordon Elliott Biotrin 93 The Rise Mount Merrion Dublin Ireland

2. SoGAT 2005 TMB VP2 HRP OD450 IgG ELISA TMB CE Marked FDA Approved l Intact Viral particles are essential for 100% detection of B19. l VP1/2 antigens expressed in E.Coli do not form particles and give 10-20% false negative IgG results. Electron Micrograph of Biotrin VP2 Capsids

3. SoGAT 2005 TMB VP2 HRP OD450 IgG ELISA TMB CE Marked FDA Approved VP2 HRP TMB OD450 IgM ELISA CE Marked FDA Approved Virus HRP TMB OD450 Antigen ELISA Research assay

4. SoGAT 2005 Vireamic donations - antibody profiles: 70 Vireamic (>10e6 IU / ml) samples collected over 12- 18 month period Biotrin examined the B19 IgG and IgM levels

5. SoGAT 2005

6. Specificity of the assay: To evaluate integrity of positive results at the low end of the range - <2.5 index In a panel of 500 blood donors, 350 (70%) were IgG positive. 18 samples (3.5%) were ‘very low pos’, index 1.1 to 2.5 These 18 samples were tested by confirmatory competitive (VP2) assay

7. SoGAT 2005 Specificity of the assay:

8. SoGAT 2005 Effectiveness of IgG donor screening: Normal donor population (Netherlands) 80% are IgG reactive Of all vireamic specimens detected by plasma PCR screen process (>10e6 IU ml), in a 12-18 month period, 0% true IgG positives (n=70) Rate of parvo IgG in normal donors is possibly at least 80 times rate in vireamic donors Typically 1 in 10,000 fresh blood product units donated are vireamic (>10e5 IU / ml) (greater in epidemic) Consider that with a 1 time IgG screen at the time of donation, the rate of vireamic donations may be better than 1 in 800,000 (Allowing for 1 false positive in this case, 1.43% are ‘apparent reactive’ rate of vireamic donations would be of the order of > 1 in 560,000)

9. SoGAT 2005 Current Sanquin Approach: The two-time testing model is clearly successful in reducing, if not eliminating, high vireamic donations. Main drawback is requirement for time gap in two-time testing One time screening using combined tests maybe possible: 1) IgG+ / IgM- donors (zero of 70 viraemics tested) 2) IgG+ / Parvo Ag- donors 3) IgG+ / PCR low donors Studies ongoing

10. SoGAT 2005 Use of antigen in Parvovirus IgG ELISAs The Biotrin assay is based on G1 (B19) VP2 Capsid: Is VP1 and VP2 required for detection of all positives? Does the Biotrin assay work for Genotype 2 (A6) and Genotype 3 (V9)? Collaboration with Prof. JP Allain, Cambridge Transfusion Medicine

11. SoGAT 2005 Servant et al, 2002 Genotype 1 (B19) VP2 Genotype 1 (B19) VP1/2 Genotype 3 (V9) VP2 Genotype 3 (V9) VP1/2 ‘CAM 1’ ‘CAM2’ ‘CAM3’ ‘CAM4’ CODE 1 CAMBRIDGE BIOTRIN CODE 2 Test 500 donors from G1 (B19) region (UK) Test 500 donors from G3 (V9) region (Ghana)

12. SoGAT 2005 138 UK Donors tested (to date) using several capsid antigens

13. SoGAT 2005 500 Ghanaian Donors: > 100 tested to date Different index value distribution compared to European populations Lower Index Values overall But no difference in results between antigens. Testing of Ghanaian Children < 5 yrs demonstrated that G3 Antisera reacts with Biotrin’s G1 antigen as well as with experimental G3 antigen. G3 derived antibodies are detected as well as G1 derived antibodies using the current assay.

14. SoGAT 2005 An Antigen-Capture EIA for the detection of Parvovirus B19 in human plasma. The first EIA for B19 Virus Detection. Employs a classic “sandwich” EIA format. Sensitivity of approximately 5 x 10 7 IU / ml. High specificity, no false-positive results in over 500 normal human plasmas tested. Capability to detect virus in the presence of high levels of anti-B19 IgM and / or IgG. Detection of variants e.g. V9

15. SoGAT 2005

16. Sanquin: Theo Cuypers Harry Bos Marco Koppelman Cambridge: JP Allain Daniel Candotti Armen Parsyan Biotrin: Shane Kerr Amanda Corcoran