Reagent for the enzymatic determination of serum total cholesterol with improved lipolytic efficiency.

We describe a sensitive method for quantifying the extent of cholesterol ester cleavage during enzymatic assay of total cholesterol in serum. Lipids are extracted from the assay mixture with chloroform/methanol (1/1 by vol), concentrated, then quantified by "high-performance" thin-layer chromatography. Although with conventional enzymatic reagents for determination of serum total cholesterol the hydrolysis of the cholesterol esters may be incomplete, a new enzymatic cholesterol reagent (Monotest Cholesterol, High Performance, Boehringer Mannheim) gives virtually complete cholesterol ester cleavage (i.e., greater than or equal to 99.5%). Use of this reagent with its improved lipolytic efficiency yields results for serum total cholesterol that are identical to those measured with a candidate reference procedure involving alkaline cholesterol ester saponification.

[1]  D. Goldstein,et al.  Glycosylated hemoglobin measured by affinity chromatography: micro-sample collection and room-temperature storage. , 1983, Clinical chemistry.

[2]  F. Mashige,et al.  Direct spectrophotometry of total bile acids in serum. , 1981, Clinical chemistry.

[3]  F. Mashige,et al.  Continuous-flow determination of bile acids in serum, and its clinical application. , 1978, Clinical chemistry.

[4]  K. Imai,et al.  A simple and sensitive assay of total serum bile acids. , 1976, Clinica chimica acta; international journal of clinical chemistry.

[5]  A. Noma,et al.  Polarographic method for rapid microdetermination of cholesterol with cholesterol esterase and cholesterol oxidase. , 1976, Clinical chemistry.

[6]  N. Hanson,et al.  Stability in urea as a measure of cardiac lactate dehydrogenase on the centrifugal analyzer. , 1974, Clinical chemistry.

[7]  P. Fu,et al.  Enzymatic determination of total serum cholesterol. , 1974, Clinical chemistry.

[8]  P. Trinder.  Determination of Glucose in Blood Using Glucose Oxidase with an Alternative Oxygen Acceptor , 1969 .

[9]  J. Folch,et al.  A simple method for the isolation and purification of total lipides from animal tissues. , 1957, The Journal of biological chemistry.

[10]  B. Brodie,et al.  A simplified method for the estimation of total cholesterol in serum and demonstration of its specificity. , 1952, The Journal of biological chemistry.

[11]  N. Hanson,et al.  Effect of protein on the determination of total bile acids in serum. , 1983, Clinical chemistry.

[12]  R. M. Tel,et al.  Incomplete Hydrolysis of Cholesteryl Esters during the Enzymatic Cholesterol Determination as Evidenced by Aqueous Cholesteryl Ester Solutions: Comparison of Six Enzymatic Procedures with the Liebermann-Burchard Method , 1980, Journal of clinical chemistry and clinical biochemistry. Zeitschrift fur klinische Chemie und klinische Biochemie.

[13]  P. Röschlau,et al.  Enzymatische Bestimmung des Gesamt-Cholesterins im Serum , 1974 .

[14]  P. Strandjord,et al.  Heat stable lactate dehydrogenase in the diagnosis of myocardial infarction. , 1962, JAMA.