Fig. 5. Characterization of FGF-2-GFP export with regard to kinetics, unspecific release and inhibition by ouabain. (A) Kinetic analysis of FGF-2-GFP export. CHO_FGF-2-GFP cells were grown in the presence of doxicyclin for the times indicated followed by FACS processing, including antibody treatment, as described in Materials and Methods. The raw data have been subjected to a weighted curve fit and are representative of two independent experiments. (B) CHO_FGF-2-GFP cells were grown in the absence of doxicyclin followed by the addition of various amounts of a supernatant derived from homogenized CHO_FGF-2-GFP cells that were grown for 48 hours in the presence of doxicyclin (lanes 1-5). Based on cell number, 0% (lane 1), 2.5% (lane 2), 5% (lane 3), 7.5% (lane 4) and 10% (lane 5) of this supernatant was added to CHO_FGF-2-GFP cells grown in the absence of doxicyclin. The PE-derived FGF-2-GFP cell surface signal was then compared with the corresponding signal of CHO_FGF-2-GFP cells grown for 48 hours in the presence of doxicyclin (set to 100%, lane 6). Lanes 7 and 8 refer to experiments under the same conditions as those in lane 6 with the exception that during the whole course of the experiment, 1 µM and 5 µM ouabain, respectively, were added to the culture medium. The data are representative of two independent experiments.