Fig. 3. Genotype confirmation of mdx^3cv spermatozoa. (A) Expression and localization of Dp71 in spermatozoa from control and mdx^3cv mice were determined by indirect immunofluorescence and western blotting using Dys2 antibody. Immunostaining in wild-type spermatozoa was localized in the postacrosomal region and in the middle piece of the flagella (A1). The absence of Dys2 immunostaining in mdx^3cv spermatozoa confirms their genotype (A2). A1' and A2' show phase-contrast images corresponding to spermatozoa from wild-type and mdx^3cv mice. (B) The presence of dystrophin products was analyzed by western blotting. In membrane fractions from wild-type spermatozoa only one protein band of 76 kDa was detected, corresponding to Dp71 (B1). In the membrane fraction of mdx^3cv spermatozoa, Dp71 was absent and no other dystrophin was found (B2).