Anti-human tissue transglutaminase 2 monoclonal antibody and application thereof

A technology of transglutaminase and antibody, applied in the direction of antibody, tissue culture, anti-enzyme immunoglobulin, etc., can solve the problems of poor tissue contact and hindering the therapeutic application of small molecule inhibitors

Inactive Publication Date: 2015-01-14
WUHAN XIEKANG BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, their poor alkylation efficacy and tissue accessibility hinder the therapeutic application of these small molecule inhibitors

Method used

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  • Anti-human tissue transglutaminase 2 monoclonal antibody and application thereof
  • Anti-human tissue transglutaminase 2 monoclonal antibody and application thereof
  • Anti-human tissue transglutaminase 2 monoclonal antibody and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1: Polypeptide Design and Synthesis

[0043] Using literature search and protein amino acid sequence analysis software, the applicant found a highly conserved amino acid sequence in the enzymatic activity region of tissue transglutaminase 2 (tGM2), the sequence of which is shown in SEQ ID NO.1. This sequence is highly conserved among mice, rats and humans. The applicant carried out the synthesis of the polypeptide according to this sequence (synthesized by NeoBiolab, USA). Part of the polypeptide was coupled with the carrier protein Keyhole Limpet Hemocyanin (KLH) (synthesized by NeoBiolab, USA), and used as an immunogen to immunize rats.

Embodiment 2

[0044] Example 2: Preparation of anti-human tissue transglutaminase 2 (tGM2) monoclonal antibody

[0045] A. Immunization of SD rats

[0046] Three SD rats aged 12 weeks were immunized with KLH-polypeptide coupled protein as antigen. 250ug antigen was dissolved in sterile phosphate buffer and heated in a water bath at 50°C for 30 minutes, then mixed with Freund's complete adjuvant to make a homogeneous oil emulsion, and the antigen oil emulsion was injected into rats the abdominal cavity. After 3 weeks, the antigen was mixed with incomplete Freund's adjuvant by the same method and injected into the abdominal cavity of rats. After another 3 weeks, the antigen was mixed with Freund's incomplete adjuvant by the same method and injected into the abdominal cavity of rats. The immunization success of KLH-polypeptide coupled protein was determined by periocular blood sampling and enzyme-linked labeling methods 2 weeks after the last immunization.

[0047] B. cell fusion

[0048]...

Embodiment 3

[0056] Example 3: Identification of the functional activity of anti-human tGM2 monoclonal antibodies

[0057] In order to detect the activity of monoclonal antibodies inhibiting tGM2, the applicant established an enzymatic reaction system. In this system, a peptide derived from human type I collagen was first modified with Biotin (Biotin-LSQQIENI), and then cross-linked to poly-lysine (poly-lysine) catalyzed by tGM2 ( image 3 ). The extent of the cross-linking reaction was monitored by avidin-HRP mediated enzymatic reaction. image 3 showed that the transamination activity of tGM2 was greatly inhibited as the concentration of anti-tGM2 antibody increased.

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Abstract

The invention discloses an anti-human tissue transglutaminase 2 monoclonal antibody and application thereof. A rat hybridoma cell strain, particularly rat anti-human tissue transglutaminase 2 (tGM2) hybridoma cell XK-1 (CCTCC NO:C201312), is established. The hybridoma cell secretes an anti-human tissue transglutaminase 2 (tGM2) monoclonal antibody; and the monoclonal antibody protein has a specific inhibition action on tissue transglutaminase 2 (tGM2), is capable of effectively inhibiting enzyme activity of mouse tGM2 and restoring activity of matrix metal proteinases (MMPs), and has the function of treating liver fibrosis.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular to a monoclonal antibody against human tissue transglutaminase 2 (tGM2), and also relates to a monoclonal antibody against human tissue transglutaminase 2 (tGM2) in the treatment of Application in mouse liver fibrosis. Background technique [0002] Liver cirrhosis is a common chronic liver disease that results from severe fibrosis of the liver, leading to a progressive loss of liver function, caused by viral infection of the liver, alcohol abuse, and other factors. Liver fibrosis also provides an environment for tumorigenesis, which not only activates tumor cells, but also promotes tumor metastasis. The results of the National Hepatitis Epidemiological Survey in 2006 showed that the carrier rate of hepatitis B surface antigen in the Chinese population was 7.18%. It is estimated that about 457 million people had been infected with hepatitis B virus, of which 93 million were HBsAg carrie...

Claims

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Application Information

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IPC IPC(8): C12N5/20C07K16/40A61K39/395A61P1/16C12R1/91
Inventor 张红军韩原平
Owner WUHAN XIEKANG BIOTECH
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