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Lipoprotein-related phospholipase A2 enzyme-linked immunosorbent assay (ELISA) kit and preparation method thereof

An enzyme-linked immunoassay detection and phospholipase technology, which is applied in measurement devices, instruments, scientific instruments, etc., can solve problems such as blanks in the application of enzyme-linked immunoassay technology, and achieves easy large-scale promotion and use, significant economic and social benefits, The effect of simplifying the operation steps

Inactive Publication Date: 2014-08-27
TIANJIN KANGERKE BIOSCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the embodiments of the present invention is to provide a simple, fast and accurate lipoprotein-associated phospholipase A2 enzyme-linked immunoassay kit and preparation method, aiming at solving the application of enzyme-linked immunoassay technology in human Lp-PLA2 immunoassay products aspect remains blank

Method used

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  • Lipoprotein-related phospholipase A2 enzyme-linked immunosorbent assay (ELISA) kit and preparation method thereof
  • Lipoprotein-related phospholipase A2 enzyme-linked immunosorbent assay (ELISA) kit and preparation method thereof
  • Lipoprotein-related phospholipase A2 enzyme-linked immunosorbent assay (ELISA) kit and preparation method thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0052] The lipoprotein-associated phospholipase A2 enzyme-linked immunoassay kit of the embodiment of the present invention includes a standard and a quality control, a coating carrier, an alkaline phosphatase enzyme marker, a chromogenic solution, an analysis buffer, and a concentrated washing solution;

[0053] in:

[0054] 1. The standard and quality control products are made by the following method: use pH7.4, 20mmol / LPBS buffer solution and fetal bovine serum to mix in a volume ratio of 3:1 to prepare a basic buffer solution, and use the basic buffer solution to dilute the lipoprotein Relevant phospholipase A2 recombinant antigens were diluted to concentrations of 1000, 500, 250, 100, 50, and 0ng / mL standard substances; lipoprotein-related phospholipase A2 recombinant antigens were diluted to a concentration of 300ng / mL high-value quality control with basic buffer and 150ng / mL low-value quality control products, the concentration measurement ranges of high and low quality...

Embodiment 2

[0066] A lipoprotein-associated phospholipase A2 ELISA kit comprises the following components:

[0067] 1. Standard product and quality control product (same as embodiment 1):

[0068] 2. coated carrier (with embodiment 1)

[0069] 3. Horseradish peroxidase marker, its preparation method is: dissolve 2mg horseradish peroxidase in 1mL deionized water, add 0.4mL50mmol / L sodium periodate solution, shake slowly at 4°C for 30min, and use pH4. 4. Dialyze with 1mmol / L sodium acetate buffer overnight, add 2mg lipoprotein-associated phospholipase A2 antibody, shake overnight at 4°C, use 400ul of 200mmol / L NaBH4 solution for reduction, dialyze with pH7.4, 20mmol / LPBS buffer overnight, and then use HPLC For secondary purification, collect protein peaks, add an equal volume of glycerol, and store at -20°C; the labeled lipoprotein-associated phospholipase A2 antibody enzyme conjugate can be diluted to the working concentration with 20% fetal bovine serum diluent, and can be stored at 4°C S...

Embodiment 3

[0079] The following provides the specific operation method for detecting lipoprotein-associated phospholipase A2 in clinical samples using the enzyme-linked immunoassay kit prepared in Example 1:

[0080] 1. Sample processing: when the sample to be tested is plasma, take 2 mL of blood with an EDTA anticoagulant tube, centrifuge at 1500 r / min for 10 minutes, and collect the supernatant; , after standing at 4°C for 30 minutes, centrifuge at 1500r / min for 10 minutes, and collect the supernatant;

[0081] 2. Sample detection: add the sample and analysis buffer to the microwell plate with lipoprotein-associated phospholipase A2 antibody on the solid phase. After 30 minutes of reaction, the lipoprotein-associated phospholipase A2 in the sample specifically binds to the solid-phase antibody. Wash away the free components, add enzyme-labeled substances and react in the dark for 30 minutes, the enzyme-labeled lipoprotein-associated phospholipase A2 antibody forms a "sandwich" complex ...

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Abstract

The invention discloses a lipoprotein-related phospholipase A2 enzyme-linked immunosorbent assay (ELISA) kit and a preparation method of the lipoprotein-related phospholipase A2 ELISA kit. The lipoprotein-related phospholipase A2 ELISA kit comprises standard substances, quality control substances, a coating carrier, an enzyme marker, a color-substrate solution, an analysis buffer solution and a concentrated cleaning solution. The preparation method of the lipoprotein-related phospholipase A2 ELISA kit includes the first step of preparing the lipoprotein-related phospholipase A2 standard substances and the lipoprotein-related phospholipase A2 quality control substances, the second step of preparing the carrier wrapped by lipoprotein-related phospholipase A2 antibodies, the third step of preparing the enzyme marker of the lipoprotein-related phospholipase A2 antibodies, the fourth step of preparing the analysis buffer solution, the fifth step of preparing the concentrated cleaning solution, and the sixth step of assembling the lipoprotein-related phospholipase A2 ELISA kit. The lipoprotein-related phospholipase A2 ELISA kit can be used for replacing other kits for quantitative detection of lipoprotein-related phospholipase A2, and is low in cost, easy and convenient to operate, high in sensitivity and capable of being widely popularized and used clinically on a large scale.

Description

technical field [0001] The invention belongs to the technical field of immunoassay and analysis, and in particular relates to a lipoprotein-associated phospholipase A2 enzyme-linked immunoassay kit and a preparation method. Background technique [0002] Atherosclerosis (atherosclerosis) is a common disease that seriously endangers human health, and has always been the focus of medical and biochemical research for a long time in the past. Because of its wide popularity and long incubation period in the human body, it often erupts in the form of fatal diseases such as ischemia, angina pectoris, myocardial infarction, stroke, coronary heart disease or heart failure, and has become the most common cause of death at this stage. [0003] Lipoprotein-associated phospholipase A2 (Lp-PLA2) is a new inflammatory enzyme in cardiovascular diseases. It belongs to PLA2 in the phospholipase family. It is a serine-dependent phospholipase that can hydrolyze and oxidize low-density lipoprotei...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/573G01N33/535
CPCG01N33/573G01N2333/916
Inventor 兰成杰
Owner TIANJIN KANGERKE BIOSCI
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