The cholinesterase activities of pectoral, biceps, and adductor muscles from selected lines of normal and genetically dystrophic chickens were investigated using spectrophotometric assays of acetylthiocholine and butyrylthiocholine hydrolysis and acryl-amide disk gel electrophoresis. It was found that predominately white-fibered muscles such as the pectoral or biceps of dystrophic chickens has 30-40 times the acetylcholinesterase activity of their normal counterparts when the data were expressed on a per protein basis. In contrast, the cholinesterase activity of the predominately red-fibered adductor muscle was little different in samples from dystrophic and normal chickens. Acryl-amide gel electrophoresis revealed that the high acetylcholinesterase activity of dystrophic pectoral and biceps muscles was associated with isoenzyme bands of cholinesterase activity characteristic of embryonic skeletal muscle. In the normal these bands disappeared from the gels before the chicks were 2 weeks of age; in dystrophic lines many of the embryonic isoenzyme bands were retained until the chickens were at least 1 year of age. The possible significance of these findings with respect to the mechanism of action of the gene causing muscular dystrophy of the chicken was discussed.