Production and characterization of antibodies against aflatoxin in laying hens

Abstract

Polyclonal antibodies against aflatoxins were obtained from egg yolks of laying hens immunized with either aflatoxin B₁ (AFB₁) or aflatoxin M₁ (AFM₁) conjugated to bovine serum albumin (BSA). An indirect enzyme‐linked immunosorbent assay (ELISA) involving the use of AFB₁‐BSA or AFM₁‐BSA conjugate and anti‐chicken IgG‐horseradish peroxidase (HRP) conjugate, was developed for monitoring antibody titers and aflatoxin analysis. Production of the antibody in hens started as early as 10 days after immunization and reached a maximum in 20 days. Competitive indirect ELISA revealed that the antibodies were most specific for AFB₁. They were cross‐reactive with other aflatoxins in the following order: B₁ (100%)>G₁ (22–28%)>B₂ (6–16%)>M₁(3–9%)>G₂ (1–4%). Anti‐AFM₁‐BSA antibodies showed a similar pattern of cross‐reaction to the anti‐AFB₁‐BSA antibodies when AFB₁‐BSA was coated to the ELISA plate. The specificity of anti‐AFM₁‐BSA to AFM₁ was demonstrated when AFM₁‐BSA was coated to the ELISA plate. Cross‐reactivity with different aflatoxins was in the following order: M₁ (531%)>B₁ (100%)>G₂ (41%)>G₁ (19%)>B₂ (3%). The sensitivity for AFB₁ analysis in the indirect ELISA was 0.05–5 ng AFB₁/assay.