Characterization of relevant membrane antigens by two-dimensional immunoblot and n-terminal sequence analysis in...

In myocarditis an antigen-specific immune response to cardiac epitopes has been demonstrated by several investigators. In 25 patients with histologically proven myocarditis, autoantibodies to cardiac tissue were observed in 65% by immunofluorescence assay. By immunoblot analysis 46% of sera reacted with cardiac tissue. These antibodies were preferentially directed against proteins with molecular weights of 43 and 67 kDa. Many of these antibodies have been demonstrated to be cytolytic to isolated cardiocytes in vitro. However, their corresponding antigens on heart muscle cells still remains unclear. To characterize the epitopes recognized by the autoantibodies, isoelectric focusing followed by SDS-PAGE was used to separate the complex mixture of proteins from human heart. Immunoblot analysis of antigens revealed proteins in the range of 30-67 kDa at isoelectric points of 6.5-8.5 to be of particular interest. Five of these proteins were identified by immunoblot analysis and two were identified by n-terminal sequence analysis (Edman Degradation) as creatine kinase and a sarcomeric isoenzyme. As creatine kinase is an intracellularly located enzyme, the pathogenetic role of the autoantibodies directed against it remains to be elucidated further.