Preparation of peroxidase conjugated monoclonal antibodies against G class of chicken immunoglobulins (IgY)

Abstract

Infections of poultry with pathogen microorganisms are relatively frequent and can cause great economic loss. Most of bacterial or virus infections can be indirectly detected by demonstration of specific antibodies. The most frequently used are indirect immunoenzyme assays where specific serum imunoglobulins of class Y (IgY), which is an avian homologue of mammals’ class G (IgG), are demonstrated. Beside the appropriate antigen in such assays, the secondary antibodies against chicken IgY, conjugated with enzyme are necessary. IgY isolated from chicken egg yolk were used for immunization of BALB/c mice and monoclonal antibodies (mAb), designated M2 were designed and conjugated with horseradish peroxidase (M2-p). MAb M2 recognized determinants on chicken, turkey, pheasant and peacock IgY. Determinant is a part of heavy chain of chicken IgY and is probably not glycosilated. For use of conjugate M2-p in direct immuno-enzyme assays pH optimum, optimal working dilution and affinity were determined. Very wide range of pH (from 3 to 11) values, where mAb M2 reacts with IgY indicates a high affinity of binding. Results obtained so far indicate that our conjugate M2-p is appropriate for use in immunoenzyme assays for detection of specific antibodies IgY in chicken, turkey, pheasant and peacock.