procleix ® wnv assay: a tma-based assay for screening blood donations for west nile virus rna
DESCRIPTION
Procleix ® WNV Assay: A TMA-based Assay for Screening Blood Donations for West Nile Virus RNA. SoGAT July 3, 2003. Jeff Linnen, Ph.D. Research and Development Gen-Probe Incorporated, San Diego, CA USA. Procleix® is a registered trademark of Chiron Corporation. TECAN. - PowerPoint PPT PresentationTRANSCRIPT
1
Procleix® WNV Assay: A TMA-based Assay for Screening Blood
Donations for West Nile Virus RNA
Jeff Linnen, Ph.D.Research and Development
Gen-Probe Incorporated,San Diego, CA USA
SoGATJuly 3, 2003
Procleix® is a registered trademark of Chiron Corporation
2
Procleix WNV Assay
• Based on Transcription Mediated Amplification (TMA)
• Uses same instrument platform as Gen-Probe’s licensed NAT blood screening assay– Procleix Semi-automated System currently used with
Procleix HIV-1/HCV Assay
TECAN Target Capture System (TCS) Luminometer
3
Specificity of the Procleix WNV Assay
• 1,680 normal blood donations were tested at Gen-Probe– 99.8% initial specificity; 100% resolved specificity– over 40,000 archived samples from 2002 high risk populations have been
tested by American Red Cross (S. Stramer)
• No cross reactivity to other blood borne viruses – Testing included HTLV, HIV-1/-2, HCV, HBV, HGV, Rubella, HAV, CMV,
EBV, HCV, Parvo B19
• Assay designed to specifically detect West Nile virus:– No cross reactivity to other flaviviruses: Dengue (1-4), Yellow Fever Virus,
and St. Louis Encephalitis virus; weak cross reactivity to Murray Valley Encephalitis virus
– Detects Kunjin virus (Australian subtype of West Nile virus)
4
Analytical Sensitivity
Percent Positive
Copy Level BBI Lineage 1Virus* (N = 10)
BBI Lineage 2Virus* (N = 20)
300 100% 100%
100 100% 100%
30 100% 100%
10 100% 100%
3 50% 65%
1 10% 5%
0.3 0% 10%
0 0% 0%*virus quantified by BBI TaqMan Assay (BBI Diagnostics, West Bridgewater, MA).
Based on data from Procleix WNV Assay kit lot manufactured at 2 million test scale
Percent Positive
Copy LevelImpath BCPLineage 1**
(N =30)50 100%
25 100%
5 100%
1 100%
0 0%**Quantitation for this panel is probably not accurate
Probit Analysis (results from BBI panels):Lineage 1: 95% detection at 4 to 14 copies/mLLineage 2: 95% detection at 4 to 8 copies/mL
5
Samples from CDC WNV Transfusion Transmission Case Investigations (2002)
Quantitation Procleix WNV AssayCase
IDCDC PCRResults
NGIPCR
Copies/mL
ARCTaqManPFU/mL
Neat 1:8 1:16 IgMSero-conversion
1 POS 7,700 16.4 ++ +++ +++ +
2 POS 3,300 13.5 ++ +++ +++ NA3 POS 3,900 7.1 ++ +++ +++ NA4 POS 1,200 2.3 ++ +++ +++ NA5 POS 31,000 1,412 ++ +++ +++ NA6 POS 6,600 13.3 ++ +++ +++ +7 POS 37,000 1,225 ++ +++ +++ +8 POS 39,000 327 ++ +++ +++ +9 POS 9,000 7.9 ++ +++ +++ +
10 NEG/POS(Hi input) 630 0.9 ++ +++ +++ +
11 NEG/POS(Hi input) 480 0.06 ++ +++ +++ +
12 NEG NEG 0.013 ++ + - - - - - +++ - - - + (EQV at index)
13 NEG POS;< 100 NEG - - - - + NT NT +
• Positive Results from testing 383 blinded samples sent to Gen-Probe from the American Red Cross• All donations implicated in transfusion transmission were detected at 1:16 dilution with TMA
6
Nationwide WNV Blood Screeningin the United States, 2003
• Testing using the Procleix WNV Assay started on June 19– Implemented nationwide on July 1– Test development which normally takes 2 to 3 years was condensed into
less than 9 months.
– Procleix WNV Assay is being used to test over 80% of the US blood supply
• Most donations are being tested in pools of 16 (some sites are testing individual donations)– reactive pools will be resolved to individual donation
• Testing will reduce the risk of WNV transfusion transmission– will provide real time surveillance of human WNV activity in North America
7
Acknowledgements• Gen-Probe WNV Assay Development Team
– (Front to back, left to right) Mike Shih, Josh Cary, Geoffrey Dennis, Janice Cline, Martha Alden, Wen Wu, Mackenzie Lewis, Michelle Cass, Amy Broulik, Jeff Linnen, Stephanie Miller
• National Heart, Lung, and Blood Institute (NHLBI) for partial funding• Susan Stramer, American Red Cross• Chiron Corporation