Determination of cholesterol with a microporous membrane chemiluminescence cell with cholesterol oxidase in solution

• Published in: Analytica chimica acta 1985, Vol.170 (2), p.199-207
• Main Authors: Malavolti, Nathan L., Pilosof, David, Nieman, Timothy A.
• Format: Article
• Language: English
• Subjects: Analytical, structural and metabolic biochemistry; Biological and medical sciences; Fundamental and applied biological sciences. Psychology; Other biological molecules; Terpenes, steroids. Hormones
• ISSN: 0003-2670; 1873-4324
• DOI: 10.1016/S0003-2670(00)81743-2

A reagent solution, containing cholesterol oxidase buffered at pH 7, is contained in a pressurized reservoir and forced through a microporous membrane at 2–5 μl min −1 into a stream flowing at 2–10 ml min −1 which contains injected slugs of cholesterol as the analyte. The hydrogen peroxide produced then reacts with luminol in pH 9.0 Tris buffer, catalyzed by horseradish peroxidase, to produce chemiluminescence, the intensity of which is related to the cholesterol concentration. The working range is 0.4–40 mg dl −1; precision is 1–4% over this range. The detection limit is 0.2 mg dl −1 or 5 μM. Sample throughput is 60 h −1, and only 0.01 unit of enzyme is consumed per sample. Blood serum samples may be analyzed for either free or total cholesterol by using standard addition and pre-treatment with Somogyi reagents for removal of reducing species.