Fig. 7. Expression and localization of chimeras in Caco-2 cells. (A) Stable clones of Caco-2 cells expressing Cav-1 (1), Cav-2 (2) or chimeras I, II, III or IV were lysed and 50 µg of each homogenate was analyzed by SDS-PAGE and immunoblotting with the polyclonal antibody against Cav-1 (N-20). Chimeras I, II, III and IV showed a slower migration because of the addition of the Myc epitope. The molecular mass markers are indicated on the left in kDa. (B) Subcellular localization of chimeras by confocal analysis of Caco-2 cells. Cells expressing Chimeras I, II, III and IV were double-labeled with rabbit polyclonal anti-Cav-1 (N20) (red) and mouse monoclonal antibody (green) against Giantin (a Golgi marker) (a,c,e,g) or Ag525 (an endogenous basolateral marker) (b,d,f,h). CH-I (I) and II (II) can be observed at the periphery of cells marked by arrows while CH-III and IV show colocalization with Giantin indicated by arrowheads. Bar, 5 µm.