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Method for measuring cholesterol in remnant-like lipoprotein

A technique for remnant-like lipoprotein and determination method, applied in the field of determination of cholesterol in remnant-like lipoprotein

Inactive Publication Date: 2009-02-25
JIMRO
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, these methods cannot necessarily be said to be sufficient in terms of the selectivity and simplicity of the enzymatic reaction.

Method used

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  • Method for measuring cholesterol in remnant-like lipoprotein
  • Method for measuring cholesterol in remnant-like lipoprotein
  • Method for measuring cholesterol in remnant-like lipoprotein

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0065] (1) Preparation of sample

[0066] The serum components were separated into remnant-like lipoprotein fraction, LDL fraction and HDL fraction, and the determination of each lipoprotein was carried out. The isolation method of this protein is as follows.

[0067] Anti-apoB-100 antibody-binding affinity chromatography columns and anti-apoA-1 antibody-binding affinity chromatography columns were prepared respectively, serum was added, and the binding fractions were recovered as LDL and HDL, respectively. Furthermore, serum was added to the aforementioned two chromatographic columns, and only the fraction that passed through was collected as a remnant-like lipoprotein fraction (RLP). Individual lipoproteins were confirmed by HPLC analysis. Furthermore, the concentration of cholesterol in each lipoprotein was adjusted to 5 mg / dl using a reagent kit for measuring total cholesterol.

[0068] (2) Preparation of measurement reagents

[0069] Reagents for measuring cholesterol...

Embodiment 2

[0089] Then, using an automatic analyzer TBA-20R (Toshiba), the amount of RLP-cholesterol in human serum was measured according to the measuring method of the present invention. NUREX R (product of the present invention) was used as a surfactant.

[0090] Sample: human serum 106 sample

[0091] table 3

[0092] Reagent 1

[0093] Good's buffer pH6.8 (PIPES, manufactured by Dojin Chemical Co., Ltd.) 50mM

[0094] HDAOS (manufactured by Dojin Chemical Co., Ltd.) 1mM

[0095] NUREX R (manufactured by Nippon Oil & Fat Co., Ltd.) 0.01%

[0096] Reagent 2

[0097] Good's buffer pH6.8 (BES, manufactured by Dojin Chemical Co., Ltd.) 50mM

[0098] 4-Aminoantipyrine (manufactured by Kanto Chemical Co., Ltd.) 3mM

[0099] Peroxidase (manufactured by Amano Enzyme Co., Ltd.) 10 units / mL

[0100] Cholesterol esterase 15 units / mL

[0101] (COE311: activity ratio 31.8, manufactured by Toyobo Co., Ltd.)

[0102] Cholesterol oxidase (manufactured by Amano Enzyme Co., Ltd.) 6 units / mL ...

Embodiment 3

[0106] The amount of RLP-cholesterol in human serum was measured by the measurement method of the present invention using an automatic analyzer 7080 (Hitachi). Furthermore, PELEX SS-L (this invention product 4) was used as surfactant, and COE311 (activity 31.8) and LP (activity ratio 512) were used as cholesterol esterase.

[0107] Sample: 30 samples of human serum

[0108] Table 4

[0109] Reagent 1

[0110] Good's buffer pH6.5 (PIPES, manufactured by Dojin Chemical Co., Ltd.) 50mM

[0111] HDAOS (manufactured by Dojin Chemical Co., Ltd.) 1mM

[0112] PELEX SS-L 0.02%

[0113] Reagent 2

[0114] Good's buffer pH6.5 (PIPES, manufactured by Dojin Chemical Co., Ltd.) 50mM

[0115] 4-Aminoantipyrine (manufactured by Kanto Chemical Co., Ltd.) 3mM

[0116] Peroxidase (manufactured by Amano Enzyme Co., Ltd.) 10 units / mL

[0117] Cholesterol esterase 10 units / mL

[0118] (COE311: activity ratio 31.8, manufactured by Toyobo Co., Ltd.)

[0119] Cholesterol esterase 20 units / m...

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Abstract

A method for measuring cholesterol in remnant-like lipoprotein in a sample, by a more convenient operation and with high sensitivity, and a reagent for the measurement are provided. A method for measuring cholesterol in remnant-like lipoprotein, the method comprising subjecting a test sample containing lipoproteins to the action of a cholesterol esterase and a cholesterol oxidase or cholesterol dehydrogenase, and measuring the hydrogen peroxide or reduced coenzyme generated by the enzymatic reaction, wherein a surfactant having a benzenesulfonic acid structure in the molecule or a polyacrylic acid-based polymeric surfactant is used.

Description

technical field [0001] The present invention relates to a method and reagent for measuring cholesterol in remnant-like lipoprotein, which is a risk factor for arteriosclerosis and the like in clinical examinations. Background technique [0002] In clinical examination, cholesterol in high-density lipoprotein (HDL) is a negative factor as a risk factor for atherosclerosis, and cholesterol in low-density lipoprotein (LDL) is a positive factor for a risk factor for atherosclerosis. are often measured in the field. [0003] In recent years, cholesterol in lipoproteins such as remnant-like lipoproteins, which are produced by lipid metabolism, has been shown to be a closer indicator than cholesterol in LDL as a risk factor for atherosclerosis. Efficient measurement of cholesterol in remnant-like lipoproteins among lipoproteins in biological samples has become a subject. [0004] As for the enzymatic assay of cholesterol in remnant lipoproteins, an assay method was reported in wh...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/44C12Q1/26C07K16/18
CPCC12Q1/46G01N2333/775G01N33/92C07K16/18C12Q1/44C12Q1/26
Inventor 荻原富美子新见学中野隆光金谷一司
Owner JIMRO