Rapid Antigen Production for Influenza Diagnostic Development
Session III – Hathaway, YI applicant
Title of Contribution: Rapid Antigen Production for Influenza Diagnostic Development
Author(s): Kyle Hathaway1, Richard Webby2, Yaeta Endo3, Julie Pavlin1, Rodney Coldren4, Shinichi Makino3, Tetsuya Ogasawara3, Tatsuya Sawasaki3, Jennifer DeBeauchamp2, Yolanda Griffith2, Ashley Webb2.
Affiliation(s): Global Emerging Infection Surveillance System, Armed Forces Research Institute of Medical Sciences1; Department of Infectious Diseases, St Jude Children’s Research Hospital2, Cell-Free Science and Technology Research Center, Ehime University3; US Army Center for Health Promotion and Preventive Medicine4
Abstract:
Influenza A viruses cause respiratory infections among humans and numerous animal species including domestic poultry and wild fowl. Continual evolution of the virus allows it to persist in both human and animal populations. For many years, diagnostic antigens to detect influenza were produced using inactivated whole virus or antigens purified from whole virus – such as bromelain cleaved hemagglutinin. While diagnostic antigens can also be produced using a variety of recombinant expression systems, the cloning, transformation, expression and purification in many systems can be a lengthy process, slowing the process of updating diagnostics in the face of a significant change in the influenza virus.
Here we present the results of cell-free expression of the hemagglutinin surface antigen from an H5N1 isolate. After expression using a wheat germ cell-free lysate milligram quantities of protein were prepared using nickel affinity purification. Both the expression and purification were accomplished in less than one week. We evaluated the diagnostic potential for use of the protein in fluorescent assays to detect serum antibodies and for the production of antibody suitable for detecting circulating virus in surveillance samples and found that this system has potential to allow for the rapid production of efficacious diagnostic antigens.
