Tissue and species specificity of antibodies produced against "native" and SDS denatured smooth muscle actin and myosin

Antisera were produced against "native" and SDS denatured forms of actin and myosin purified from chicken gizzard to investigate the following questions:

1. Is SDS denaturation prior to immunization necessary to elicit antisera against a protein previously classified as "nonantigenic" in the native state (actin)?

2. Will SDS denaturation of actin and myosin prior to immunization alter the tissue and species specificity of the corresponding antisera?

The detection of precipitating antibodies and the tissue and species specificity of the antisera were analyzed by Ouchterlony double immunodiffusion and indirect immunofluorescent staining of fixed frozen sections. Chicken gizzard, chicken cardiac, and chicken skeletal tissues were used to test the tissue specificity of the antisera while pigeon gizzard, turtle gut, guinea pig gut, and toad gut were used to test the species specificity.

Since two out of three rabbits injected with "native" actin produced precipitating antibodies, it was shown that SDs denaturation prior to immunization is not necessary for antibody production.

Differences in the tissue and species specificity of antisera produced against "native" and SDS denatured forms of the two proteins were not observed. All anti-actins were found to be equally tissue and species nonspecific. In addition, all anti-myosins were found to be tissue specific for smooth muscle but species nonspecific.

Since SDs denaturation unfolds globular proteins into more extended polypeptide chains, it can be concluded from these studies that some antigenic determinants governing species cross-reactivity must be formed by linear amino acid sequences (sequential determinants) rather than conformationally induced juxtapositions of nonsequential amino acids (conformational determinants).