Localization of pro-uPA-activating activity by subcellular fractionation. THP-1 and U937 cells were fractionated by sucrose density gradient centrifugation and the fractions assayed for pro-uPA-activating activity. Data are represented as absorbance due to hydrolysis of the uPA-specific chromogenic peptide substrate S-2444 for THP-1 cells (closed bars) and U937 cells (open bars). Plasma membrane fractions, identified by biotinylation of cell surface proteins (fractions 2 to 5), are shown by the horizontal bar. Matriptase was subsequently detected in these fractions by Western blotting (upper panel). The minor peak of activity in fractions 12 and 13 did not contain detectable levels of matriptase.