Fig. 2. Uptake and processing of extracellular heparanase pro-enzyme. (A) Western blot analysis of cell-associated heparanase. Human foreskin fibroblasts were incubated with 10 µg/ml recombinant 65 kDa heparanase for 30, 60 and 120 minutes. At each time point, the cells were lysed and subjected to western blot analysis using anti-heparanase mAb 130. (B-D) Subcellular localization of cell-associated heparanase. Human foreskin fibroblasts were incubated with 10 µg/ml recombinant 65 kDa heparanase for 30 (B), 60 (C) and 120 (D) minutes. The cells were fixed, permeabilized and subjected to immunofluorescent staining with anti-heparanase mAb 130. Note the accumulation of heparanase in granules at 60 (C) and 120 (D) minutes. Bar, 20 µm.