Chemistry: analytical and immunological testing – Lipids – triglycerides – cholesterol – or lipoproteins
Patent
1996-11-19
1999-03-30
Wallenhorst, Maureen M.
Chemistry: analytical and immunological testing
Lipids, triglycerides, cholesterol, or lipoproteins
436 13, 436166, 435 11, 435 19, 435 28, 422 61, 2524081, G01N 3392
Patent
active
058888276
DESCRIPTION:
BRIEF SUMMARY
TECHNICAL FIELD
The present invention relates to a method for the determination of cholesterol in low-density lipoprotein (LDL) or very low-density lipoprotein (VLDL) (hereinafter referred to as LDL cholesterol or VLDL cholesterol) which is important with respect to lipid metabolism in the field of clinical diagnosis.
BACKGROUND ART
LDL is considered to have the function to supply cholesterol to peripheral cells and to be a direct factor in occurrence of various types of arteriosclerosis such as coronary arteriosclerosis. It is known that LDL level in blood is useful as an indicator of arteriosclerosis. The relationship between VLDL which is rich in triglycerides (TG) and arteriosclerosis has also been noted. Currently, the determination of LDL cholesterol is carried out by the ultracentrifugation method, the electrophoretic method, the conversion method, etc., and that of VLDL cholesterol is carried out by the ultracentrifugation method, the electrophoretic method, etc. In the ultracentrifugation method, which is employed as a basic method, LDL or VLDL is separated by the difference in specific gravity using an ultracentrifuge for separation, and the amount However, this method is defective in accuracy, simplicity, economic efficiency, etc. In the electrophoretic method, LDL or VLDL is separated by using a cellulose acetate membrane or agarose gel as a support, and the (Rinsho Kensa), 29, 1344 (1985)!. This method is defective in simplicity, economic efficiency, etc. In the conversion method, the amount of LDL Chem.,18, 499 (1972)!. cholesterol)-(Amount of triglycerides)/5 type of hyperlipemia, etc., and so this method is defective in simplicity, accuracy, applicability to the analysis of a large number of samples, etc. As described above, conventional methods for the determination of LDL cholesterol or VLDL cholesterol are not suitable for the analysis of a large number of samples, the rapid analysis, and the analysis with an autoanalyzer which is widely used in the field of clinical testing. Further, in these methods, manual errors are liable to occur, for example, when the amount of the LDL fraction separated is determined using a measuring pipette. However, if a blood serum sample is directly added to a reagent containing cholesterol esterase and cholesterol oxidase without fractionation of LDL or VLDL, the resultant test system is not different from a system for the determination of total cholesterol, and LDL cholesterol or VLDL cholesterol cannot be specifically determined.
DISCLOSURE OF THE INVENTION
The present inventors have determined the amount of cholesterol in high-density lipoprotein (HDL), LDL, VLDL, and chylomicron (CM), each of which had been fractionated through ultracentrifugation, using a reagent for the determination of cholesterol containing a sugar compound and/or a protein solubilizing agent, and found that these lipoproteins differ in reactivity to the reagent based on the combinations of the sugar compound and/or the protein solubilizing agent, which leads to the difference in the reactivity of cholesterol in HDL, LDL cholesterol, VLDL cholesterol, and cholesterol in CM. This finding has led to the completion of the present invention.
The present invention relates to a method for the determination of LDL cholesterol or VLDL cholesterol in a sample, which comprises determining the amount of LDL cholesterol or VLDL cholesterol in the sample in the presence of a sugar compound and/or a protein solubilizing agent. In this method, a divalent metal salt may be added to the determination system in order to improve the specificity.
The present invention also provides a reagent for the determination of cholesterol in LDL or VLDL, which contains a sugar compound and/or a protein solubilizing agent; and a reagent for the determination of cholesterol in LDL or VLDL, which is a kit composed of a sugar compound and a protein solubilizing agent.
As the sugar compound, glucose derivatives are preferably used. Examples of the glucose derivative include compounds represented by general form
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Irie Tetsumi
Kayahara Norihiko
Shutoh Eiko
Sugiuchi Hiroyuki
Tatano Toshio
Kyowa Medex Co., Ltd.
Wallenhorst Maureen M.
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