Hybridoma cell capable of secreting anti-cardiac troponin I monoclonal antibodies and applications thereof

A technology of cardiac troponin and hybridoma cells, which is applied in the field of immunoassay of cardiac troponin I, can solve the problems of inconvenient use, large investment, narrow linear range, etc.

Active Publication Date: 2013-06-26
FAPON BIOTECH INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the traditional ELISA method has many shortcomings such as long measurement period, relatively low sensitivity, and narrow linear range; chemiluminescence and enzyme-linked fluorescence analysis methods have high sensitivity and good specificity, but they require special equipment, are inconvenient to use, and require a lot of investment. bigger

Method used

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  • Hybridoma cell capable of secreting anti-cardiac troponin I monoclonal antibodies and applications thereof
  • Hybridoma cell capable of secreting anti-cardiac troponin I monoclonal antibodies and applications thereof
  • Hybridoma cell capable of secreting anti-cardiac troponin I monoclonal antibodies and applications thereof

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Experimental program
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Effect test

Embodiment 1

[0027] Embodiment 1 The establishment of hybridoma cells and the preparation of anti-cardiac troponin I monoclonal antibody

[0028] 1. Antigen Immunization

[0029] Human recombinant cardiac troponin I antigen (1.5mg / ml, produced by Shenzhen Faipeng Biological Co., Ltd., product number AG-CTNI-HP0005) and an equal volume of Freund's complete adjuvant (appearance: amber cell suspension; Components: Paraffin Oil85%, Mannide Monooleate15%, Mycobacterium smegmatis1mg / ml) are mixed to obtain an oily emulsion. The emulsion was subcutaneously administered to the back site of BALB / c mice (Guangzhou Provincial Experimental Animal Center, 6-week-old female, 5) at a dose of 0.2ml, and the abdominal cavity was boosted immunization 14 days after the first immunization (antigen and equal amount of Fu Mixed with incomplete adjuvant), after boosting immunization to four injections, the tail blood was collected for titer detection, and the titer reached the fusion requirement.

[0030] Thre...

Embodiment 2

[0067] The preparation of embodiment 2 cardiac troponin I colloidal gold detection test paper

[0068] 1. Preparation of Nitrocellulose Membrane

[0069] Preparation of coating buffer: 6% methanol, 0.01M pH7.2 PBS buffer as coating buffer, filtered through 0.22μm membrane, set at 4°C for later use, valid for one week. 1000ml 6% Methanol 0.01M pH7.2PBS Buffer Formula: NaCL8g, KCL0.2g, NaCL 2 HPO 4 12H 2 O2.9g, KH 2 PO 4 0.2g, methanol 60ml, distilled deionized water to 1000ml.

[0070] Preparation of nitrocellulose membrane: Dilute the anti-cardiac troponin I rabbit polyclonal antibody (manufactured by Shenzhen Faipeng Biological Co., Ltd., product number PAB-CTNI-AP0002) to 1-5 mg / ml with coating buffer, adjust Machine, marked as T line, which is the detection line, the T line is close to the end of the gold standard pad, about 5mm away from the end of the gold standard pad; the goat anti-mouse IgG antibody (produced by Shenzhen Feipeng Biological Co., Ltd. , Cat. No. B...

Embodiment 3

[0094] Embodiment 3 detects the kit of cardiac troponin I

[0095] 1. The kit for rapid detection of cardiac troponin I comprises: the detection test paper and sample diluent made in Example 2.

[0096] Among them, the sample diluent is 8% NaCl solution. Preparation method: 80gNaCl, add distilled water to make up to 1000ml.

[0097] 2. Detection of cardiac troponin I by colloidal gold method

[0098] (1) Pipette 120 μl of collected human serum or plasma directly into the sample hole of the test paper card, and wait for 15 minutes to observe the results.

[0099] (2) Judgment of results: When the test strip shows a red quality control line visible to the naked eye, but no red detection line visible to the naked eye, the result is judged as negative; when the test strip shows a red quality control line visible to the naked eye, it also appears Visible red test line, the result is judged as positive. The darker the detection line, the higher the cardiac troponin I level of th...

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Abstract

The invention relates to a hybridoma cell capable of secreting anti-cardiac troponin I monoclonal antibodies, which can be rapidly and conveniently applied to hcTnI detection; and the invention also relates to a monoclonal antibody secreted by the hybridoma cell and the application thereof in the field of detecting cardiac troponin I. The hybridoma cell is preserved in the Preservation Center of Wuhan University at Mountain Lojia, Wuchang, Wuhan, Hubei (namely, China Center for Type Culture Collection), and the preservation number is CCTCC No: C201317. The secretion yield of the hybridoma cell is high, anti-cardiac troponin I monoclonal antibodies secreted by the hybridoma cell have the advantages of high affinity and high specificity and the like, and can be widely used in the detection field of anti-cardiac troponin I preparation, such as the fields of detection reagent or detection equipment preparation, and the like, and compared with traditional detection methods, the detection method has significant advantages in the aspects of specificity, sensitivity and detection rate and the like.

Description

technical field [0001] The invention relates to the field of cardiac troponin I immune detection, in particular to a hybridoma cell capable of secreting anti-cardiac troponin I monoclonal antibody and its application. Background technique [0002] Before the 1980s, the World Health Organization (WHO) has always regarded myocardial enzyme activity as one of the diagnostic criteria for acute myocardial infarction (AMI). In the late 1980s, researchers discovered that the sensitivity and specificity of troponin (Tn) was higher than that of phosphocreatine kinase (CK), phosphocreatine kinase isoenzyme (CK-MB), lactate dehydrogenation enzymes and biomarkers such as aspartate aminotransferase. Cardiac troponin I (cTnI) exists only in the myocardium and is a marker of cardiomyocytes. Its abnormal changes can affect the diastolic and systolic functions of the heart, and can be used to diagnose myocardial necrosis and judge myocardial injury. One of the most specific markers is reco...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/20C07K16/18G01N33/68G01N33/577G01N33/532C12R1/91
Inventor 杨耿周朱碧银林育佳刘莉莉
Owner FAPON BIOTECH INC
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