Immunocytochemical localization of the mammalian voltage-dependent sodium channel using polyclonal antibodies against the purified protein.
Antibodies were raised in rabbits against the purified voltage-dependent sodium channel from rat skeletal muscle sarcolemma. The resultant antiserum reacted with the purified channel in a solid-phase radioimmunoassay and precipitated the sodium channel from a crude mixture of solubilized membrane proteins. Crude membrane proteins separated according to size under nondenaturing conditions by chromatography on Sepharose CL-6B contained a single peak of immunoreactivity that coincided with the native channel. On immunoblots of sarcolemmal membrane proteins, the antiserum reacted predominantly with a diffuse high molecular weight band that was comparable in migratory characteristics to the large glycoprotein subunit of the purified channel. Using immunocytochemical techniques, binding of this polyclonal antiserum was localized to the surface membrane of rat skeletal muscle. This staining was specifically blocked by pre-incubation of the antiserum with the purified channel protein. The antiserum also stained the surface membrane of rat cardiac muscle and the nodes of Ranvier in rat peripheral nerve. Species cross-reactivity was seen with mouse, human, and guinea pig skeletal muscle while chicken, rabbit, and frog muscle was not stained. The antiserum also reacted with the surface membranes of fetal rat muscle in tissue culture. These results indicate that sodium channels in adult mammalian skeletal muscle, cardiac muscle, and peripheral nerve and in fetal muscle in culture all share common antigenic determinants. The antiserum should prove useful for topographical studies of sodium channel distribution in these tissues.
D000818:Animals; D000906:Antibodies / Q000276:immunology*; D004591:Electrophoresis, Polyacrylamide Gel; D004594:Electrophysiology; D006651:Histocytochemistry; D007120:Immunochemistry; D007473:Ion Channels / Q000378:metabolism* / Q000502:physiology; D008565:Membrane Proteins / Q000276:immunology; D009124:Muscle Proteins / Q000276:immunology*; D011817:Rabbits; D051381:Rats; D012508:Sarcolemma / Q000276:immunology / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*; D014018:Tissue Distribution
D000906:Antibodies; D007473:Ion Channels; D008565:Membrane Proteins; D009124:Muscle Proteins; D012964:Sodium