Figure 4.

Binding of fluorochrome-labeled GM₁ to immobilized FGF-2. (A) Increasing concentrations of BODIPY-12-GM₁ were incubated for 5 min at room temperature into wells coated with 20 μg/ml native (•) or heat-denatured (▪) FGF-2. At the end of incubation, the BODIPY-12-GM₁ bound to the immobilized growth factor was extracted and measured with a spectrofluorimeter. In panel B, binding data were analyzed by the Scatchard plot procedure. They are representative of three independent experiments that gave similar results. (C) Aliquots (50 μl) containing 1 nmol of BODIPY-12-GM₁ were incubated for 5 min at room temperature into FGF-2-coated wells in the presence of 100 nmol of asialo-GM₁ (□) or of increasing concentrations of unlabeled GM₁ (•). At the end of incubation, the amount of BODIPY-12-GM₁ bound to the immobilized growth factor was extracted, measured, and compared with the amount of BODIPY-12-GM₁ bound to immobilized FGF-2 in the absence of any competitor. Each point is the mean of two determinations in duplicate. SEM never exceeded 5% of the mean value.