Figure 5.

The levels of shed active matriptase are roughly correlated with the levels of matriptase expression. The conditioned buffer (shed fraction) was collected from Ramos cells following acid-induced matriptase zymogen activation. The shed fraction was then subjected to immunodepletion using the activated matriptase mAb M69 bound to Sepharose beads. The shed fraction (A: Acid sup; B: lanes 1), the activated matriptase-depleted shed fraction (A: M69 Del; B: lanes 2), and the control shed fraction (A: PBS sup) were analysed for tryptic activity using a synthetic fluorogenic substrate (A), for matriptase species by immunoblot using the total matriptase mAb (B: left panel, Total MTP), and gelatinolytic activity by gelatine zymography (B: right panel, Gelatine Zymog.). Half a million cells of each of the seven-different neoplastic B-cell lines were transiently exposed to a pH 6.0 buffer. The shed fraction was collected and analysed for the tryptic activity. The rate (RFU/min) of the tryptic activity is presented per 2.5 × 105 cells (C) or normalised to the level of matriptase expressed by these seven lines (D). RFU stands for relative fluorescent unit. The immunodepletion studies and matriptase enzymatic activity assays were conducted at least two times, and representative data are presented.