Determn. of enzymes which deactivate bleomycin - by measuring DNA-polymerase inhibition in presence and absence of test sample

摘要

Determination of the bleomycin-deactivating enzyme activity of biological liquids comprises first measuring the degree of DNA-polymerase inhibition caused by a given quantity of bleomycin (I) under controlled conditions then comparing this with the corresp. inhibition in the presence of test liq. A reagent for carrying out the test is an aq. soln. of 0.01-1mm each of deoxy-ATP, deoxy-CTP and deoxy-GTP; 120um tritium-labelled deoxythymidine triphosphate; 0.01-1 mg/ml DNA; 0.01-0.2m buffer of pH 7-7.6; 5-100 mm MgCl2; 5-100mm dithiothreitol; 5-50 ug/ml serum albumin; 5-20 U/ml DNA polymerase and opt. 0.1-2 mm mercaptoethanol. (I) is a cytostatic agent for treating laminar epithelial carcinoma, but is deactivated by enzymes present in many tissues. For correct choice of doage this inhibition needs to be known. The present method is rapid, sensitive and suitable for routine use.