Isolation, Production, and Characterization of Serum Immunoglobulin M (IgM) of Indian Major Carp Cirrhinus mrigal (MRIGAL)

Abstract

A method for the isolation of immunoglobulin M (IgM) in Indian major carp Cirrhinus mrigal (mrigal) serum to produce polyclonal antibodies is described in the present study. The purified immunoglobulins (IgM) were isolated from the serum of mrigal (Cirrhinus mrigal) by the bovine serum albumin (BSA)-CL affinity column purification method, and the IgM was used to produce a polyclonal rabbit anti-mrigal IgM antiserum. The IgM preparations were employed in the characterization of mrigal serum immunoglobulin. Reduced mrigal IgM on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was shown to consist of two subunits, compatible with heavy and light chains. A single heavy chain at approximately 90 kDa and variant of light chain 30 kDa were found. The dominant form of nonreduced IgM had a MW of approximately 900 kDa, suggesting a tetrameric structure based on estimated molecular weights, the relative protein content, and the reactivity with anti-mrigal IgM antisera, was obtained. The antisera were characterized as to specificity and reactivity by means of the enzyme linked immuno-sorbent assay (ELISA) and western blotting method. The information on the structure and character of immunoglobulin of fishes is essential in health management. The study described here investigates the possibility of using the serological techniques to assess the reactivity of antibody with the anti-mrigal IgM antisera.