Purification of Influenza virus A (H1N1) recombinant Hemagglutinin (HA1) and polyclonal antibody production

By changing expression conditions such as IPTG (Isopropyl β-D-1-thiogalactopyranoside) concentration, time and temperature of incubation, the expression conditions for HA1 were optimized. The total cell protein harvested and purified by nickel affinity chromatography. All above mentioned experiments monitored by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Results: The efficiency of HA1 recombinant protein was high, equal to 400-600 mg/ml of cell lysate. The polyclonal antibody was prepared by immunizing the rabbits using recombinant HA1 with Freund’s adjuvant according to standard protocols. Efficiency of the antiserum evaluated by enzyme linked immunosorbent assay (ELISA). Determination of antibody level in the collected antiserum using serum-based ELISA showed that the specific antibody has risen well through the immunization schedule. Conclusion: Our data shows that this polyclonal antibody has potential to be produced in rabbit. It will also be used in the future in influenza diagnosis as well as in other immunological applications such as western blot analyses, immunocytochemistry, and immunohistochemistry. Abstract