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Antigen binding part of anti-human Tim-3 monoclonal antibody

A technology of monoclonal antibody and antigen, which is applied in the field of the above-mentioned monoclonal antibody or its antigen-binding part, monoclonal antibody or its antigen-binding part, and can solve the problems of unmarketed Tim-3 antibody, etc.

Active Publication Date: 2015-05-06
GENERAL HOSPITAL OF PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although studies have shown that neutralizing antibodies to Tim-3 play a very good role in intervening in certain disease models, there are currently no commercially available Tim-3 antibodies with neutralizing activity

Method used

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  • Antigen binding part of anti-human Tim-3 monoclonal antibody
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  • Antigen binding part of anti-human Tim-3 monoclonal antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0105] Example 1 Preparation of anti-human Tim-3 monoclonal antibody L3B

[0106] 1. Materials

[0107] Freund's complete adjuvant and Freund's incomplete adjuvant, chromogenic reagent TMB: product of Sigma company; 20% fetal bovine serum: product of Beijing Yuanheng Shengma Biotechnology Institute; serum-free RPMI 1640: product of Gibco company; SP2 / 0 cells: imported from ATCC and preserved by the Institute of Basic Medical Sciences, Academy of Military Medical Sciences; BALB / c mice were provided by the Experimental Animal Center, Academy of Military Medical Sciences; recombinant human Tim-3 (rhTim-3) was prepared by the Institute of Basic Medical Sciences, Academy of Military Medical Sciences. The rest of the reagents were purchased from the market.

[0108] 2. Method

[0109] Monoclonal antibodies were obtained using standard in vivo immunization methods and PEG fusion methods. For detailed methods, see: Ed Harlow et al., Antibodies A Laboratory Manual, Cold Spring Harbo...

Embodiment 2

[0117] Example 2 Fishing of human Tim-3 monoclonal antibody L3B light chain variable region and heavy chain variable region genes

[0118] 1. Materials

[0119] (1) Upstream and downstream primers for the light chain variable region

[0120] The sequences of the light chain upstream primers MuLC5, MuLC6 and MuLC7 are shown in the sequence listing SEQ ID NO: 11, SEQ ID NO: 12 and SEQ ID NO: 13 respectively; the light chain downstream primer MuCK is shown in the sequence listing SEQ ID NO: 14 , the concentration of primers used in PCR amplification was 10 pmol / μL.

[0121] (2) Primers for the upstream and downstream of the heavy chain variable region

[0122] For the heavy chain upstream primers MuHC5, MuHC6, MuHC7 and MuHC8, the sequences are shown in the sequence listing SEQ ID NO: 15, SEQ ID NO 16, SEQ ID NO: 17 and SEQ ID NO: 18; for the heavy chain downstream primer MuIgG2a, see SEQ ID NO: 15 in the sequence listing ID NO: 19, the concentration of primers used in PCR amp...

Embodiment 3

[0133] Example 3 Expression and Purification of Monoclonal Antibody L3B

[0134] 1. Materials

[0135] Protein A Sepharose CL 4B column Protein column: product of Beijing Benyuan Zhengyang Biotechnology Co., Ltd.; pcDNA3.1 plasmid: Invitrogen; sources of other reagents and materials are the same as in Example 1.

[0136] 2. Method

[0137] The monoclonal antibody L3B light chain variable region gene obtained in Example 2, and the light chain constant region gene were loaded into the pcDNA3.1 plasmid to obtain a vector containing the monoclonal antibody L3B light chain gene; the monoclonal antibody obtained in Example 2 The L3B heavy chain variable region gene and the heavy chain constant region gene are loaded into the pcDNA3.1 plasmid to obtain a vector containing the L3B heavy chain gene of the monoclonal antibody. The vector containing the light chain gene of the monoclonal antibody L3B and the vector of the heavy chain gene are co-transfected into mammalian cells for exp...

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Abstract

The invention discloses an antigen binding part of an anti-human Tim-3 monoclonal antibody L3B. The antigen binding part of the monoclonal antibody L3B can be specifically combined with human Tim-3 protein. The antigen binding part of the anti-human Tim-3 monoclonal antibody can be used for preparing a kit for detecting human Tim-3, and the kit is relatively high in sensitivity and can detect out human Tim-3 with relatively low concentration. The invention further provides a related hybridoma cell strain, a separated nucleic acid molecule as well as drug composition and an immunodetection tool comprising the monoclonal antibody.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and relates to an anti-human Tim-3 monoclonal antibody or an antigen-binding part thereof. The present invention also relates to the use of the above-mentioned monoclonal antibody or its antigen-binding part, including preparing human Tim-3 protein immunodetection tools and preparing pharmaceutical preparations for treating diseases related to abnormal expression of Tim-3. Background technique [0002] In 2001, McIntire and others discovered a new gene family, which structurally contains immunoglobulin V region and mucin region, so it was named T cell immunoglobulin-and mucin-domain-containing molecules (T cell immunoglobulin-and mucin-domain-containing molecules , Tim) family. Tim protein is specifically expressed on Th1 or Th2 cells, which is a new important marker for distinguishing Th1 and Th2, and plays an important role in T cell differentiation, T cell effector function, autoimmune...

Claims

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Application Information

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IPC IPC(8): C07K16/28C12N15/13A61K39/395A61P37/02A61P11/06A61P31/18A61P31/14A61P31/20G01N33/68G01N33/577
Inventor 康春燕肖燕刘长庭方向群曲宝林李洪霞苏茂生赵永生蔡博宁蒋兴伟韩根成
Owner GENERAL HOSPITAL OF PLA
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