Immunological identification of avian monomeric and polymeric immunoglobulin M and immunoglobulin A after fractionation on sodium dodecylsulfate pore gradient polyacrylamide gels

Naturally existing variants of Immunoglobulin M (IgM) and Immunoglobulin A (IgA) in the serum of immunologically competent, as well as immunologically defective UM-B19 chickens can be detected without time consuming purification. The different molecular forms of IgM and IgA were separated by gel filtration. Pentameric IgM and dimeric IgA were well separated from the 7S-peak that contained monomeric IgM and IgA. A method for immunological identification of the protein components is described. Pore gradient gel electrophoresis was combined with antigen-antibody crossed electrophoresis in horizontal agarose slabs. The principle of the method is that IgM and IgA in their different molecular forms in the mixtures to be assayed after gel filtration are first separated with high resolution by sodium dodecylsulfate (SDS) polyacrylamide gel electrophoresis using a pore gradient of 3 to 15%. In the second step, a slab of polyacrylamide gel with the separated proteins in transferred onto agarose layers containing appropriate anti- mu- and anti-alpha antiserum and SDS in a concentration of .01%. The technique is easy to perform and gives reproducible results. In chicken serum, the monomeric state of IgM (184,000 d), along with the pentameric state (920,000 d) were identified. Serum IgA exists in dimeric (340,000 d) and in monomeric (170,000 d) states. No differences between the commercial Leghorn and dysgammaglobulinemic and normogammaglobulinemic lines were found.