Production and purification of polyclonal anti-chicken IgY immunoglobulins in Goats and conjugation with Horseradish Peroxidase

Abstract

Antibodies are very important components in medical diagnostics and various biochemical assays for diagnosis of the diseases. IgY is an immunoglobulin largely present in Avians. Anti-chicken IgY immunoglobulins conjugated with gold nanoparticles are used to diagnose the chicken infectious diseases by lateral flow assay (LFA) and enzyme linked immunosorbent assay (ELISA) immunodiagnostic tests. The main aim of this study is the production of antichicken IgY antibodies in goats, purification of antiantibody and conjugation with horse radish peroxidase (HRP) enzyme. The antibody was purified using protein G sepharose affinity column and the purity of the antibody was confirmed by SDS-PAGE analysis and by spectrophotometer. The purified anti IgY antibody was also evaluated by ELISA. The optimum dilution of prepared HRP conjugated anti IgY was 1:9600. The sensitivity and specificity of the antibody were evaluated and have no cross reactivity with other IgG antibodies. This study showed that protein G affinity purification could be best useful method for purification of polyclonal IgG antibodies.