Development of a high-throughput cell based 384-well influenza A quantification assay for interpandemic and highly pathogenic avian strains
- 1
- 2
- 3
- 4
- 5
- 6
Abstract
Influenza remains a world wide health threat, thus the need for a high-throughput and robust assay to quantify both seasonal and avian in-fluenza A strains. Therefore, a 384-well plate format was developed for the median tissue culture infectious dose assay (TCID50) utilizing the detection of nucleoprotein by an in situ en-zyme linked immunosorbent assay (ELISA) which was optimized for sensitivity in this assay. Highly pathogenic avian influenza, A/Vietnam/ 1203/04 (H5N1), and interpandemic strains, A/ New Caledonia/20/99 (H1N1) and A/Brisbane/ 10/07 (H3N2), were quantified using this high- throughput assay. Each 384-well plate can be used to analyze ten viral samples in quadrupli-cate, eight dilutions per sample, including all necessary assay controls. The results obtained from 384-well plates were comparable to tradi-tional 96-well plates and also demonstrate re-peatability, intermediate precision, and assay linearity. Further, the use of 384-well plates in-creased the throughput of sample analysis and the precision and accuracy of the resulting titer.
- (2009) Influenza statistics. Roche.
- (2009) WHO confirmed human cases of avian influenza a (H5N1). World Health Organization Department Co- mmunicable Disease Surveillance and Response.
- Tewari, D., Zellers, C., Acland, H. and Pedersen, J.C. (2007) Automated extraction of avian influenza virus for rapid detection using real-time RT-PCR. Journal of Clinical Virology, 40, 142-145.
- Lee, W.-M., Grindle, K., Pappas, T., Marshall, D.J., Moser, M.J., Beaty, E.L., Shult, P.A., Prudent, J.R. and Gern, J.E. (2007) High-throughput, sensitive, and accu-rate multiplex PCR-microsphere flow cytometry system for large-scale comprehensive detection of respiratory viruses. Journal of Clinical Microbiology, 45, 2626-2634.
- Nemoto, H., Komoriya, T. and Kohno, H. (2007) Devel-opment of latex turbidimetric immunoassay for rapid and sensitive detection of influenza virus. Rinsho Biseibutshu Jinsoku Shindan Kenkyukai Shi., 18, 117-126.
- Severson, W. E., McDowell, M., Ananthan, S., Chung, D.H., Rasmussen, L., Sosa, M.I., White, E.L., Noah, J. and Jonsson, C.B. (10-1-2008) High-throughput screen-ing of a 100,000-compound library for inhibitors of in-fluenza a virus (H3N2). Journal of Biomolecular Screening, 13, 879-887.
- Wang, S.Y., Su, C.Y., Lin, M., Huang, S.Y., Huang, W. I., Wang, C.C., Wu, Y.T., Cheng, T. J., Yu, H.M., Ren, C.T., Wu, C.Y., Wong, C.H. and Cheng, Y.S. (2-11-2009) HA-pseudotyped retroviral vectors for influenza an- tagonist screening. Journal of Biomolecular Screening, 1087-0571, 0833-0786.
- Kapteyn, J.C., Porre, A.M., de Rond, E.J.P., Hessels, W.B., Tijms, M.A., Kessen, H., Slotboom, A.M.E., Oerlemans, M.A., Smit, D., van der Linden, J., Schoen, P. and Thus, J.L.G. (2-25-2009) HPLC-based quantification of haema- gglutinin in the production of egg- and MDCK cell- de-rived influenza virus seasonal and pandemic vaccines. Vaccine, 27, 1468-1477.
- Wagaman, P.C., Leong, M.A. and Simmen, K.A. (2002) Development of a novel influenza a antiviral assay. Journal of Virological Methods, 105, 105-114.
- Mo, C., Yamagata, R., Pan, A., Reddy, J., Hazari, N. and Duke, G. (2008) Development of a high-throughput Ala-mar blue assay for the determination of influenza virus infectious dose, serum antivirus neutralization titer and virus ca/ts phenotype. Journal of Virological Methods, 150, 63-69.
- Aguero, M., San Miguel, E., Sanchez, A., Gomez-Tejedor, C. and Jimenez-Clavero, M. (2007) A fully automated procedure for the high-throughput detection of avian in-fluenza virus by real-time reverse transcriptionpoly-merase chain reaction. Avian Diseases, 51, 235-241.