Fig. 3. Direct interaction of dystrobrevin and kinesin heavy chain in rat brain. (A) The anti-dystrobrevin antibody (Clone 23) co-immunoprecipitates kinesin and dystrobrevin isoforms. Pre-cleared rat cerebellum lysate (input), mouse IgG control immunoprecipitate (IP: control IgG) and anti-dystrobrevin immunoprecipitate (IP: dystrobrevin) were separated by SDS-PAGE, and transferred to nitrocellulose. The blot was probed with anti-kinesin (Clone H2) and the polyclonal anti-ß-dystrobrevin antibody, and revealed by ECL. Besides detecting ß-dystrobrevin (59 kDa), the polyclonal anti-ß-dystrobrevin antibody cross-reacts with -dystrobrevin isoforms. (B) The anti-kinesin antibody (Clone H2) co-immunoprecipitates dystrobrevin isoforms and kinesin. Rat cerebellum lysate (input), anti-kinesin immunoprecipitate (IP: kinesin) and mouse IgG control immunoprecipitate (IP: control IgG), were separated by SDS-PAGE and transferred to nitrocellulose. The blot was probed with anti-dystrobrevin (Clone 23) and anti-kinesin (Clone H2) antibodies, and revealed by ECL. The anti-dystrobrevin antibody (Clone 23) is reactive to all dystrobrevin isoforms. In the anti-kinesin immunoprecipitate (IP: kinesin) a band corresponding to -dystrobrevin 1 (78 kDa) is clearly visible. The 59 and 55 kDa bands of dystrobrevin isoforms (see input) overlap with the co-migrating 55 kDa IgG heavy chain band (see IP: control IgG). Molecular masses of size markers are indicated on the right-hand side (in kDa). WB, western blot.