Method of producing plasminogen activator

#The present invention is directed to a method for obtaining plasminogen activator, the enzyme which converts plasminogen to plasmin.

#Urokinase, the well known plasminogen activator produced by renal cells and urinary tract is excreted in the urine and has been the subject of much interest for possible use as a medicament against sudden fibrinolytic disturbances such as thromboembolism.

#One major set of difficulties hindering wide spread employment of urokinase are the problems encountered in the recovery thereof. Human urine, the first recognized source of urokinase contains a low proportion of the enzyme, and therefore human urine for source material purposes offers extreme logistic problems as well as concern that some of the urine donors might also be excreting viruses or other detrimental substances in the urine.

#For further understanding of the practice of this invention reference is made to the following examples.

#Specimens of numerous cell types (see table 1) propagated in Falcon flasks were resuspended in Eagles basal medium (described in Grand Island's Biological Catalogue) and seeded into the wells of a multi-welled trays (Limbro) at 200,000 cells/well in 1 ml medium. Cultures were incubated overnight at 36.degree. C. Resulting confluent cultures were washed and overlayed with 199 medium. Medium for one set of cultures contained 0.25% w/v skimmed milk (for detection of proteinase). Medium for another set of cultures, contained both 0.25% skimmed milk and human serum at 1:30 as a source of plasminogen (for detection of activator). Following further incubation at 36.degree. C., the following results were obtained:

#TABLE I ______________________________________ Cell Type Plasminogen Pro- Designation Source Activator tease ______________________________________ Diploid strains MRC-5 PDL 31 Human embryo lung + - MRC-5 PDL 22 (diploid strains) + - HEL-Ki PDL 3 + - WI-38 PDL 28 + - IMR-90 PDL 26 + - Continuous lines HEp2 Human careinoma - - L 929 Mouse - - Hep.M Hepatoma-human - - Vero Monkey kidney - - BHK 21 Baby hamster kidney + - LLC-MK.sub.2 Monkey Kidney + .+-. Primary/Secondary Cultures Monkey Kidney Rhesus (2.degree.) + + Human embryo kidney Primary + + Mouse fibroblasts Mouse embryos (2.degree.) + - ______________________________________ +Indicates clearing of milk with absence of TCA precipitable protein.