Automated Fluorimetric Determination of Cellular Cholesterol

Abstract

We developed a completely automated fluorimetric method for the determination of cellular cholesterol, consisting of enzymatic hydrolysis of cholesteryl ester to free cholesterol and enzymatic oxidation of free cholesterol in the presence of an indicator substrate to produce a fluorescent product. For control preparations of monocytes, the mean detection limit was 2.57 mumol/5 x 10(5) cells and the mean within-batch coefficients of variation were 9.30, 6.00 and 3.73% at mean cholesterol concentrations of 1.94, 9.05 and 12.49 mumol/5 x 10(5) cells, respectively. The results correlated well with those obtained by gas-liquid chromatography.