Plasminogen activator in cultured Lewis lung carcinoma cells measured by chromogenic substrate assay

A chromogenic substrate assay for the plasminogen activator (PA) activity of Lewis lung carcinoma cells has been developed. The cells were incubated with plasminogen, the activation of which to plasmin was measured by the amidolysis of the chromogenic substrate S-2251. This was routinely performed as a 4h serum-free assay, but a variation lasting 24 h, in medium supplemented with plasminogen-free inhibitor-reduced serum, produced similar results. The assay also detected PA released into the medium. PA activity was proportional to cell density, and the assay was non-toxic to the cells. Assays were performed on cultures derived from primary and metastatic tumours. Host cells were effectively eliminated from such cultures but, because of an initial phase of tumour-cell death, PA assays were not carried out until cultures became established. No consistent difference was detected between PA levels in primary and metastatic cultures. However, these cultures were shown to be atypical of the parent tumour; they grew slowly when reinjected at the primary site, and their metastatic potential was impaired.