Figure 4

Effect of culture conditions on p53 in human foreskin fibroblasts. (A) Monolayer cells (50,000/35 mm dish) were cultured for 24 hr in FBS as indicated, followed by immunoblotting for p53 and GAPDH (loaded with equal cell number). The image of the p53 immunoblot has been contrast-enhanced. (B) Monolayer cells at the indicated plating density (35 mm dish) were grown for 24 hr, followed by immunoblotting for p53 and GAPDH (equal volumes); A = A-431 control lysate. (C) Phase images from panel B; bar = 20 μm. (D) Relative p53 densitometry from 3 experiments of the type in panel B. (E) Attached (att) collagen matrices containing fibroblasts at the indicated starting concentration were incubated for 24 hr prior to being released (rel) for the indicated minutes, followed by immunoblotting for p53 and GAPDH (loaded with equal cell number); control lysate A-431 on p53 gel only. (F) Similar experiment to panel E, except over a narrower cell concentration range; a = attached; r = released. (G) Relative p53 densitometry from 3 experiments of the type in panel F.