Highly sensitive cholesterol assay with enzymatic cycling applied to measurement of remnant lipoprotein-cholesterol in serum.

Abstract:

BACKGROUND:Remnant lipoprotein-cholesterol (RLP-C) concentrations in sera of healthy individuals are very low (0.080-0.437 mmol/L), making conventional cholesterol methods poorly suited to this purpose. We have developed a highly sensitive cholesterol assay (CD method) and applied it to the RLP-C assay. METHODS:The CD shuttled cholesterol reversibly between reduced and oxidized forms in the presence of thio-NAD and NADH. The production rate of thio-NADH correlated with the cholesterol concentration and was measured by the absorbance at 404/500 nm. This CD method was combined with an immunoaffinity separation procedure with specific monoclonal antibodies to apolipoprotein (apo) A1 and apo B-100 and used for RLP-C assay. Results were compared with a RLP-C method that uses cholesterol oxidase, peroxidase, and chromogenic substrate. RESULTS:The CD method could detect 0.10 x 10(-3) mmol/L cholesterol and was at least 5 times more sensitive than the conventional enzymatic method. Within- and between-day imprecision (as CVs) of the RLP-C assay with the CD method was <4%. Regression analysis of RLP-C assays with the new (y) and conventional (x) cholesterol methods yielded: y = 1.02x - 0.008 mmol/L (S(y/x) = 0.0065 mmol/L; r = 0.997; n = 297). CONCLUSIONS:Serum RLP-C can be measured by the CD method. The CD method may be useful for other assays that require sensitive cholesterol measurements in biological materials.

journal_name

Clin Chem

journal_title

Clinical chemistry

authors

Kishi K,Ochiai K,Ohta Y,Uemura Y,Kanatani K,Nakajima K,Nakamura M

subject

Has Abstract

pub_date

2002-05-01 00:00:00

pages

737-41

issue

5

eissn

0009-9147

issn

1530-8561

journal_volume

48

pub_type

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