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Metformin monoclonal antibody hybridoma cell line and application thereof

A technology of hybridoma cell lines and monoclonal antibodies, which is applied in the preparation methods of animal/human proteins and peptides, serum albumin, etc., can solve the problems of high detection limit, complexity, and low sample purity requirements, and achieve good detection sensitivity , good specific effect

Active Publication Date: 2019-12-27
JIANGNAN UNIV
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Problems solved by technology

High-performance liquid chromatography is used for detection. The detection method is cumbersome and complicated, and the detection limit is too high. In order to protect the interests of consumers, it is necessary to establish an efficient and rapid detection method for MET. ) is simple in pretreatment, low in cost, can realize rapid detection of a large number of samples, and does not require high purity of samples during detection

Method used

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  • Metformin monoclonal antibody hybridoma cell line and application thereof

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Embodiment 1

[0023] Example 1 Preparation of hybridoma cell line Tcf 1A6

[0024] (1) Synthesis of complete antigen: Weigh 1.8 mg of MET derivative 4cpMET (the molar ratio of 4cpMET to bovine serum albumin (BSA) is 30:1), dissolve it in 300 μL N,N-dimethylformamide (DMF) , stirred and reacted for 10 minutes, after fully dissolving, 1.3 mg of N-hydroxysuccinimide (NHS) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) were successively added 2.1 mg, activated at room temperature for 4-6 hours, as solution A. Take 6mg of BSA, dissolve it with 2mL 0.01M carbonate buffer solution (CB, pH=9.0) (referred to as solution B), then slowly add solution A to solution B drop by drop, and perform coupling reaction at room temperature overnight; then use 0.01 M PBS solution was dialyzed to remove the unreacted small molecule hapten to obtain the complete antigen 4cpMET-BSA, which was identified by ultraviolet absorption scanning method;

[0025] (2) Animal immunization: After mixing...

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Abstract

The invention discloses a metformin monoclonal antibody hybridoma cell line and application thereof, and belongs to the field of food safety immunoassay. The prepared metformin monoclonal antibody hybridoma cell line Tcf 1A6 is deposited in the general microbiology center of the china microbial species collection management committee, the deposit date is March 7, 2019, and the preservation numberis CGMCC No. 17399. According to the metformin monoclonal antibody hybridoma cell line and application thereof, the complete antigen of the metformin is mixed and emulsified with an equivalent amountof Freund adjuvant, and BALB / c mice is immunized through multi-point subcutaneous injections of neck and back parts to screen hybrid cells after fusion of two kinds of cells; and then cells are screened through indirect competitive enzyme-linked immunosorbent assay and subcloned for three times, and the monoclonal antibody hybridoma cell line Tcf 1A6 is finally obtained. The provided monoclonal antibody secreted by the cell line Tcf 1A6 has better specificity and detection sensitivity to MET(the IC<50> value is 1ng / mL), the residual amount of MET in chicken, chicken liver, and feed can be detected, raw materials are provided for the immunoassay of the MET residues in food, and the practical applicational value is obtained.

Description

technical field [0001] The invention relates to a metformin monoclonal antibody hybridoma cell line and application thereof, and belongs to the field of food safety immunoassay. Background technique [0002] Metformin (Metformin, MET) is a hypoglycemic drug that can inhibit type Ⅱ diabetes mellitus, especially obese type Ⅱ diabetes mellitus, which is ineffective in simple diet control and physical exercise. The combination of metformin and insulin can reduce the dosage of insulin and prevent the occurrence of hypoglycemia. It can be used in combination with sulfonylurea hypoglycemic agents and has a synergistic effect. Metformin can promote the utilization of glucose by surrounding tissue cells (muscle, etc.) and inhibit hepatic gluconeogenesis, thereby reducing the output of liver glucose and inhibiting the uptake of glucose by intestinal wall cells. The effect of metformin is different from that of insulin, that is, this product has no effect on promoting fat synthesis. ...

Claims

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Application Information

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IPC IPC(8): C12N5/20C07K16/44G01N33/577G01N33/531C07K14/765C07K1/34C12R1/91
CPCC07K14/765C07K16/44G01N33/531G01N33/577G01N2430/00
Inventor 胥传来许晓昕匡华徐丽广孙茂忠刘丽强吴晓玲宋珊珊胡拥明
Owner JIANGNAN UNIV
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