A method for preparing skeletal muscle fiber basal laminae

Abstract

Previous attempts to prepare skeletal muscle basal laminae (BL) for ultrastructural analyses have been hampered by difficulties in successfully removing skeletal muscle proteins and cellular debris from BL tubes. In the present study we describe a two phase method which results in an acellular muscle preparation, the BL of which are examined by light, transmission electron, and scanning electron microscopy. In the first phase, excised rat extensor digitorum longus muscles are subjected to x-radiation and then soaked in Marcaine to inhibit muscle regeneration and to destroy peripheral muscle fibers. The muscles are then grafted back into their original sites and allowed to remain in place 7-14 days to allow for maximal removal of degenerating muscle tissue with minimal scar tissue formation. In the second phase, the muscle grafts are subjected sequentially to EDTA, triton X-100, DNAase, and sodium deoxycholate to remove phagocytizing cells and associated degenerating muscle tissue. These procedures result in translucent, acellular muscle grafts which show numerous empty tubes of BL backed by endomysial collagenous fibers. These preparations should be useful for morphological analyses of isolated muscle BL and for possible in vitro studies by which the biological activity of muscle BL can be examined.