Sensitive assay for cholesterol in biological membranes reveals membrane-specific differences in kinetics of cholesterol oxidase
✍ Scribed by Crockett, Elizabeth L. ;Hazel, Jeffrey R.
- Publisher
- John Wiley and Sons
- Year
- 1995
- Tongue
- English
- Weight
- 606 KB
- Volume
- 271
- Category
- Article
- ISSN
- 0022-104X
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✦ Synopsis
Abstract
Quantification of cholesterol in biological membranes from a variety of sources is an important step toward understanding cholesterol's roles in membranes function. We extend to biological membranes the fluorometric/enzymatic approach (cholesterol oxidase) to measures cholesterol, originally described for whole cells (Heider and Boyett [1978] J. Lipid Res., 19: 514–518; Gamble et al. [1978] J. Lipid Res., 19: 1068–1070) and serum (Huang et al. [1975] Clin Chem., 21: 1605–1608). This method has a detection limit of 0.3 μg cholesterol. As revealed by comparison with high‐performance liquid chromatography, the fluorometric/enzymatic method with biological membranes is accurate (within 4% and 8% for intestinal and hepatic plasma membranes, respectively). The assay may be completed within 3 to 4 hours and requires neither lipid extraction nor chromatographic techniques. Kinetics of the cholesterol oxidase reaction are membrane‐specific, and first‐order rate constants (k) are positively correlated with membrane order. © 1995 Wiley‐Liss, Inc.