Production and application of a monoclonal antibody based peroxidase conjugate for detection of chicken IgG antibodies in ELISA

Abstract

Viral infections are the cause of great economic losses in the poultry industry. Development of appropriate reagents and serological diagnostic kits will help to control these infections. The aim of this study is to prepare a peroxidase labeled anti-chicken IgG using a MAb (5B8) against chicken IgG, for detection of chicken antibodies in ELISA. Hybridoma cells producing the MAb 5B8 were cultured in RPMI 1640 medium and the MAb was purified from the cells supernatant using a sepharose matrix column, sensitized with chicken IgG. The purified MAb was labeled with Horseradish peroxidase (HRP) by periodate treatment. The peroxidase labeled MAb was compared with a commercial polyclonal product for detection of chicken antibodies against avian influenza virus nucleoprotein in ELISA. Therefore, type A recombinant nucleoprotein influenza virus was used as the antigen and chicken sera prepared from healthy and influenza virus infected chickens were used as primary antibodies. The results showed that there is a strong and direct correlation (r =0.972) between the optical densities of a commercial anti-chicken IgG and the prepared conjugate. In conclusion, the conjugated MAb is appropriate for the development of serological diagnostic tests for poultry infections.

Keywords

• chicken
• igg
• monoclonal antibody
• peroxidase conjugate