Creatine kinase isoenzyme MB detection kit and preparation method thereof

A technology of creatine kinase and detection reagent, applied in the field of fluorescence immunochromatography detection, can solve the problems of dependence, complicated operation, long detection time, etc.

Pending Publication Date: 2020-11-03
上海艾瑞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

At present, the quantitative diagnosis of CK-MB isoenzymes is mainly based on large-scale chemiluminescence instruments. These methods are costly, complicated to operate, and take a long time to detect. Most of them rely on foreign imports. The high cost leads to high testing prices in hospitals. no less

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  • Creatine kinase isoenzyme MB detection kit and preparation method thereof
  • Creatine kinase isoenzyme MB detection kit and preparation method thereof
  • Creatine kinase isoenzyme MB detection kit and preparation method thereof

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preparation example Construction

[0124] A specific preparation method of the detection kit of the present invention comprises the steps of:

[0125] a. Mix the mouse creatine kinase isozyme MB monoclonal antibody-fluorescent microsphere conjugate with buffer A, wherein the mass concentration of fluorescent microspheres is 0.1-0.5 mg / mL;

[0126] b. Quantitatively and accurately divide the above mixed solution into the six-three-hole sampling cups, the filling volume is 2.5-10 μL per prefabricated cup, and place it in a vacuum drying tank for 0.5-3 hours to vacuum dry. The sample cup plate is punched into an independent sample cup containing the mouse creatine kinase isozyme MB monoclonal antibody-fluorescent microsphere conjugate;

[0127] c. Assemble the sample pad, nitrocellulose membrane coated with mouse-derived creatine kinase isozyme CK-MB polyclonal antibody, goat anti-mouse IgG polyclonal antibody, and the absorbent pad on the PVC bottom plate, and cut into fixed wide reagent cards; and

[0128] d. ...

Embodiment 1

[0138] Solution preparation during the preparation of mouse creatine kinase isozyme MB monoclonal antibody-fluorescent microsphere conjugate:

[0139] a. Preparation of blocking solution: Weigh 0.0992g disodium hydrogen phosphate and 0.0358g sodium dihydrogen phosphate and dissolve them in 80mL of water, shake and mix well. After the disodium hydrogen phosphate and sodium dihydrogen phosphate are completely dissolved, add 2.5g trehalose , 2.5g glycerin and 0.1g sodium azide, shake and mix well, adjust the pH to 8.5 with hydrochloric acid and sodium hydroxide, and finally set the volume to 100mL at room temperature.

[0140] B. configuration buffer B, the formula of buffer B: the molar concentration of PB (phosphate buffer saline) is 10mM, and the mass concentration of trehalose is 2.5%, and the mass concentration of glycerol is 2.5%, and the massfraction of sodium azide is 0.1%, pH adjusted to 8.5.

Embodiment 2

[0142] Preparation method of creatine kinase isoenzyme CK-MB immunofluorescence chromatography reagent card

[0143] 1. Replace the solution of fluorescent microspheres (carboxyl fluorescent latex microspheres) with a labeling buffer solution of pH=5.5-7.0 by centrifugation and washing;

[0144] 2. Add 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) to activate fluorescent latex microspheres. After the reaction is completed, Centrifuge to remove the supernatant, and wash again;

[0145] 3. Add mouse-derived creatine kinase isozyme MB monoclonal antibody and activated fluorescent latex microspheres to react. After the reaction is completed, centrifuge to remove the supernatant, and wash once again to obtain mouse-derived creatine kinase isozyme monoclonal antibody - Fluorescent microsphere conjugates;

[0146] 4. Block the obtained mouse creatine kinase isoenzyme monoclonal antibody-fluorescent microsphere conjugate with the ab...

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Abstract

The invention provides a creatine kinase isoenzyme MB detection kit and a preparation method thereof. Specifically, the kit comprises a creatine kinase isoenzyme MB detection reagent card, and the reagent card comprises a sample pre-adding part and a test strip; wherein the pre-sampling part comprises a sampling container and a creatine kinase isozyme MB monoclonal antibody-fluorescent microsphereconjugate which is pre-added into the sampling container; the test strip comprises a test strip bottom plate, and a sample loading part, a detection part and a liquid absorption part which are arranged on the test strip bottom plate, wherein the detection part comprises a chromatography matrix, a detection band and a quality control band, wherein the detection band and the quality control band are respectively arranged, the detection band is coated with a creatine kinase isozyme CK-MB polyclonal antibody, and the quality control band is coated with an IgG polyclonal antibody. The kit is simple to operate, low in cost, good in repeatability and high in accuracy.

Description

technical field [0001] The invention relates to the field of fluorescence immunochromatography detection, in particular to a kit for quantitatively detecting CK-MB in clinical samples by using fluorescence immunochromatography. Background technique [0002] Acute myocardial infarction (AMI) has become one of the major diseases that threaten human life. At present, 17 million people die from cardiovascular diseases in the world every year, and more than half of them die from acute myocardial infarction. The enzymes in cardiomyocytes during AMI, such as Creatine kinase (CK) is highly active in serum and can be widely used in the diagnosis of AMI. [0003] Creatine kinase (CK) is a dimer composed of brain-type subunit B and muscle-type subunit M. The molecular weight of each subunit is about 43kDa. The M subunit and B subunit have the same primary structure The origin is up to 80%, and it will form 3 kinds of dimers in the advanced structure, including CK-MM, CK-BB and CK-MB t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/573G01N33/577G01N33/558G01N33/58
CPCG01N33/573G01N33/577G01N33/582G01N33/558G01N2333/9123G01N2800/324
Inventor 陆亮倪晓涛肖琨林叶顾晓慧
Owner 上海艾瑞德生物科技有限公司
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