Chemiluminescent enzymatic assay for cholesterol in serum using lucigenin

A new highly-sensitive assay for cholesterol in serum is described. This assay utilizes chemiluminescence (CL) generated by the reaction of lucigenin and hydrogen peroxide. The latter is produced by the cholesterol/cholesterol oxidase system. Since lucigenin reacts not only with hydrogen peroxide but also with many reductants in serum to give CL, means for suppressing such undesirable CL have been investigated. This was successfully achieved by the addition of copper(II) ions, guanidine hydrochloride and Triton X-100, and accurate adjustment of the pH to 11.75–11.9 with sodium phosphate buffer. Under the optimal conditions the detection limit (2σ) for cholesterol was 1 mg 1−1, and gave results that correlated linearly with a conventional assay up to 2.5 g 1−1.