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Clinical Chemistry 24: 108-114, 1978;
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Clinical Chemistry, Vol 24, 108-114, Copyright © 1978 by American Association for Clinical Chemistry

Automated enzymic measurement of total cholesterol in serum

GS Rautela and RJ Liedtke

We describe a completely automated enzymic system for measuring total cholesterol in serum. All reagents are contained in an analytical test pack and the test is performed on Du Pont's Automatic Clinical Analyzer (aca), which mixes the sample (20 microliter) and reagents and performs the necessary absorbance measurements and calculations. In the procedure, cholesterol oxidase oxidizes free cholesterol. The oxidation step produces cholest-4-en-3-one and hydrochloride peroxidesterase hydrolyzes cholesterol esters and cholesterol in direct proportion to the amount of cholesterol present. N,N-Diethylaniline hydrochloride and 4-aminoantipyrine react with the hydrogen peroxide to produce a quinoneimine dye (lambda max = 553 nm). Interacting reagents have been optimized simultaneously (cooptimization) utilizing response surface designs coupled with computer analysis of the data. Reagent efficiency is high and analytical performance reliable.


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