A rapid assay method of collagenase activity using 14C-labeled soluble collagen as substrate.
A rapid assay method for vertebrate collagenase (EC 3.4.24.3) activity has been developed using 14C-labeled soluble collagen as substrate. The method is based on the incubation of collagen with enzyme in the presence of glucose to prevent collagen fibril formation followed by selective extraction of the enzyme digestion products into dioxane at a final concentration of 50%. The rate of reaction was about 10 times higher than that obtained by the conventional method using reconstituted collagen fibrils as substrate and the relationship between enzyme activity and concentration was linear over a wider range. When the method was applied to the assay of human granulocyte collagenase, the results showed good correlation with those obtained by the conventional gel method.
D000818:Animals; D002250:Carbon Radioisotopes; D003094:Collagen; D004589:Electrophoresis, Disc; D006098:Granulocytes / Q000201:enzymology; D006801:Humans; D007553:Isotope Labeling; D007700:Kinetics; D008722:Methods; D003012:Microbial Collagenase / Q000032:analysis*; D011489:Protein Denaturation; D012867:Skin / Q000201:enzymology; D012995:Solubility; D013696:Temperature
D002250:Carbon Radioisotopes; D003094:Collagen; D003012:Microbial Collagenase