Enzymatic assay of total cholesterol in serum or plasma by amperometric measurement of rate of oxygen depletion following saponification.
A method for serum or plasma cholesterol assay involving amperometric measurement of the rate of oxygen depletion in the cholesterol oxidase-catalyzed oxidation of cholesterol is described. The hydrolysis of the serum cholesterol esters is accomplished by saponification of 50 mul of sample with 0.2 ml of ethanolic KOH (1.0 mol/1) containing 0.5% Triton X-100 for 5 min at 75 degrees C. The rate of oxygen consumption in a 25-mul aliquot of this is measured with a Clark electrode in a Beckman Glucose Analyzer and the assay takes about one minute after incubation; results are read digitally on the instrument. The analyzer cell contains 1 ml of 1 M phosphate buffer, pH 7.4, with 100 mg sodium cholate/100 ml and 0.1-0.2 U cholesterol oxidase.
D002021:Buffers; D055598:Chemical Phenomena; D002621:Chemistry; D002784:Cholesterol / Q000097:blood*; D002793:Cholic Acids / Q000494:pharmacology; D004563:Electrochemistry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D006913:Hydroxysteroid Dehydrogenases / Q000097:blood; D066298:In Vitro Techniques; D008722:Methods; D010101:Oxygen Consumption / Q000187:drug effects; D011092:Polyethylene Glycols / Q000494:pharmacology
D002021:Buffers; D002793:Cholic Acids; D011092:Polyethylene Glycols; D002784:Cholesterol; D006913:Hydroxysteroid Dehydrogenases