抄録
An electrophoretic protocol previously used for separation of rat myosin heavy chain (MHC) isoforms was slightly modified to improve the separation of human MHC isoforms in both large and mini gel systems. The addition of reducing agents (β-mercaptoethanol or dithiothreitol) to the top running buffer (TRB) radically improved separated MHC isoform resolution and intensity of electrophoretic runs lasting longer than 5 h. In mini gel systems, the MHC isoforms could be separated in as little as 5 h. The improved resolution of bands with the inclusion of reducing agents to the TRB facilitated identification of clear boundaries for densitometric quantification of relative MHC isoform content, particularly for MHC IIa and MHC IIx. No significant effect of these reducing agents added to the TRB was observed for runs lasting only 100 min. Thus, the inclusion of reducing agents to the TRB is essential for long electrophoretic runs, usually when separating large molecular mass proteins.
