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Kit for quantitatively detecting heart-type fatty acid binding protein, and preparation method thereof

A fatty acid combination and quantitative detection technology, applied in the field of biomedicine, can solve the problems of poor specificity and low sensitivity, and achieve the effect of high specificity, high sensitivity, and guaranteed sensitivity

Inactive Publication Date: 2018-08-24
浙江艾明德生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The invention provides a kit and preparation method for quantitative detection of heart-shaped fatty acid binding protein, which overcomes the shortcomings of low sensitivity and poor specificity in the prior art

Method used

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  • Kit for quantitatively detecting heart-type fatty acid binding protein, and preparation method thereof
  • Kit for quantitatively detecting heart-type fatty acid binding protein, and preparation method thereof
  • Kit for quantitatively detecting heart-type fatty acid binding protein, and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Example 1 Preparation of H-FABP Acridine Lipid Labeled Monoclonal Antibody

[0054] 1) Weigh an appropriate amount of H-FABP monoclonal antibody, and use 0.05mol / L pH 9.5 CB to adjust to 1mg / mL;

[0055] 2) Acridine lipid and antibody are calculated according to 1:15 mole ratio, dissolved in DMF, mixed, and reacted at room temperature for 30 minutes;

[0056] 3) The reaction solution was transferred to a dialysis bag (molecular weight cut-off 8000-12000), and dialyzed with 0.01M PBS with pH 7.2 for 24 hours;

[0057] 4) Add an equal volume of glycerin and 0.1% PC-300 by volume.

Embodiment 2

[0058] Example 2 Preparation of the H-FABP quantitative assay kit of the present invention

[0059] 1. Preparation of acridine lipid antibody

[0060] 1. Acridine-labeled monoclonal antibody diluent Buffer A:

[0061] Buffer A: 0.01-0.1M PBS pH 7.2, 0.1%-1%BSA, 0.2% PC-300

[0062] 2. Select the concentration of acridine-labeled monoclonal antibody:

[0063] The working concentration of acridinium-labeled monoclonal antibody is greater than 0.41μg / mL.

[0064] 2. Preparation of H-FABP calibrator

[0065] Dilute the H-FABP antigen with 0.01M PBS pH 7.2, 0.5% bovine serum albumin (BSA) diluent, and prepare five kinds of calibrator with standard concentration of 1, 5, 25, 100, 500ng / mL.

[0066] 3. Preparation of acridine-labeled antibody:

[0067] 1) Weigh an appropriate amount of H-FABP monoclonal antibody, and use 0.05mol / L pH 9.5 CB to adjust to 1mg / mL;

[0068] 2) Acridine lipid and antibody are calculated according to 1:15 moles, dissolved in DMF, mixed, and reacted a...

Embodiment 3

[0094] Example 3 Buffer Buffer

[0095] The buffer is an important component of the present invention, which is a buffer for diluting biotinylated antibodies, antibodies labeled with acridine derivatives, and magnetic particles coupled with anti-biotin antibodies, using a combination of BSA, trehalose, and casein The formula improves the non-specific physical adsorption of immunoglobulin by magnetic particles, adding sheep immunoglobulin G (sheep IgG) and mouse IgG to further reduce the specific binding, reduce the HAMA effect, and improve the analytical sensitivity of the reagent.

[0096] Buffer A: 0.01-0.1M PBS pH 7.2, mass ratio 0.1%-1%BSA, volume ratio 0.2% PC-300;

[0097] Buffer B: use 0.01-0.1M 4-hydroxyethylpiperazineethanesulfonic acid (HEPES), 150mM NaCl, mass ratio 0.1%-1%BSA, mass ratio 0.1%-5% trehalose, mass ratio 0.5%-1 % casein sodium salt, rabbit IgG 5mg / mL, sheep IgG 3mg / mL, mouse IgG 1 mg / mL, volume ratio 0.2% PC-300, pH 7.2.

[0098] In one embodiment, t...

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Abstract

The invention provides a kit for quantitatively detecting a heart-type fatty acid binding protein. The kit comprises a heart-type fatty acid binding protein calibrator, a magnetic particle coupled with an anti-biotin antibody, an acridine derivative-labeled heart-type fatty acid binding protein monoclonal antibody, a biotin-labeled heart-type fatty acid binding protein monoclonal antibody, a chemiluminescent substrate and a quality control substance. A preparation method of the kit comprises the following steps: preparing the calibrator from a pure heart-type fatty acid binding protein raw material; preparing the acridine derivative-labeled antibody; preparing a biotinylated antibody; coupling a magnetic particle with the anti-biotin antibody; sub-packaging the calibrator, a marker mixturesolution and the chemiluminescent substrate; and performing assembling to obtain the finished product. The kit has the advantages of high sensitivity, good specificity, high accuracy of a quantitative detection result, low use cost, and easiness in promotion and application.

Description

technical field [0001] The invention relates to the field of biomedicine, in particular to a kit and a preparation method for quantitatively detecting heart-type fatty acid binding protein. Background technique [0002] Heart-type fatty acid-binding protein (h-FABP) is a novel small cytosolic protein enriched in the heart that is highly cardiac-specific (ie, predominantly expressed in cardiac tissue). Following the onset of myocardial ischemic injury, h-FABP can be detected in the blood as early as 1 to 3 hours after the onset of chest pain, peaks at 6 to 8 hours and plasma levels return to normal within 24 to 30 hours. Cardiac fatty acid binding cytoplasmic protein consists of 132 amino acids and has a molecular weight of 15 kDa. The h-FABP gene is located on chromosome I. It is one of the most abundant proteins in the heart. h-FABP binds two fatty acid molecules and participates in the transport of fatty acyl-CoA, active in the oxidation process to generate energy in th...

Claims

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Application Information

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IPC IPC(8): G01N33/68
CPCG01N33/68
Inventor 陈志强胡国富朱炎
Owner 浙江艾明德生物科技有限公司
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