To investigate the precise localization of cytoplasmic gamma actin in skeletal muscle and the relationship to dystrophin molecules, we designed an antibody against the N-terminal peptide of cytoplasmic gamma actin. Western blot analysis using SDS-PAGE and isoelectric focusing (IEF) gel revealed that the antibody reacted only with the actin isoforms having gamma motility, confirming that the antibody is specific to the cytoplasmic (nonmuscle) gamma actin. Immunohistochemical analysis of the skeletal muscle of the adult mouse revealed a dotlike staining pattern of the antibody in transverse sections and a striated staining pattern in longitudinal sections. The double immunostaining technique revealed the colocalization of cytoplasmic gamma actin with a-actinin, implying the localization of the actin on the Z-disc. Contrary to previous findings (1), we did not detect the colocalization of cytochrome oxidase, a mitochondria. marker, with this actin.