Fig. 4. Presence of transiently expressed GFP-3a60 and GFP-3a66 in nuclear speckles and CBs. (A-D) The localization of transiently expressed GFP-3a60 and GFP-3a66 was analyzed after fixation of HeLa cells in cold methanol. The left panels show the fluorescence of GFP-3a60 (A and B) and GFP-3a66 (C and D). Images in the middle panels were obtained after immunostaining with anti-SC35 (A and C) or anti-p80-coilin (B and D) and rhodamine-coupled secondary antibodies. The right panels represent computer-generated overlays of the images shown on the left and in the middle. Colocalization of GFP-3a60 and GFP-3a66 with p80-coilin in CBs (indicated by arrows) appears in yellow in (B) and (D). CBs stained by GFP-3a60 and GFP-3a66 but not by SC35 appear green in the overlays in (A) and (C). Bar, 10 µm.