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Monoclonal antibodies against hla-g isoform molecules hla-g5 and hla-g6 and uses thereof

A technology of HLA-G5 and HLA-G6, applied in the direction of anti-receptor/cell surface antigen/cell surface determinant immunoglobulin, anti-animal/human immunoglobulin, application, etc., can solve the problems of incorrect expression and emergence false positives etc.

Active Publication Date: 2022-03-08
TAIZHOU ENZE MEDICAL CENT GROUP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, antibodies 2A12 and 5A6G7 can also recognize other unknown non-HLA-G5 and HLA-G6 molecules with a molecular weight in the range of 36-175kDa. There may be cross-reactions in the application of immunohistochemistry, false positives, and incorrect HLA -G5 and HLA-G6 molecule expression
[0007] Therefore, in the analysis of the expression of HLA-G molecules and the judgment of clinical significance, the existing antibodies for detecting HLA-G molecules are not all satisfactory, and there is an urgent need for more specific and new monoclonal antibodies against HLA-G molecules

Method used

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  • Monoclonal antibodies against hla-g isoform molecules hla-g5 and hla-g6 and uses thereof
  • Monoclonal antibodies against hla-g isoform molecules hla-g5 and hla-g6 and uses thereof
  • Monoclonal antibodies against hla-g isoform molecules hla-g5 and hla-g6 and uses thereof

Examples

Experimental program
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Embodiment 21

[0046] Example 2.1. The antibody (9Y5G6) was used to detect HLA-G5 and HLA-G6 molecular recognition specificity by immunoblotting.

[0047] After HLA-G isoform standard protein HLA-G1, HLA-G2, HLA-G3, HLA-G4, HLA-G5, HLA-G6 and HLA-G7 molecular standard protein electrotransfer membrane, with 5% skimmed milk powder at room temperature Blocked for 4h, washed with 0.2% TBS (Teween-20 PBS). Add the antibody (9Y5G6) to detect its recognition specificity, incubate overnight at 4°C, and wash; add HRP-labeled rabbit anti-mouse IgG antibody, incubate at room temperature for 30min, wash with Dako REAL TM EnVision TM The detection system (DAKO) was incubated for 1-3min. The results show that: the results show that the antibody (9Y5G6) can specifically recognize HLA-G5 and HLA-G6 standard proteins, and there is no cross-reaction with other HLA-G isoform molecules ( Figure 5 ).

Embodiment 22

[0048] Example 2.2. The antibody (9Y5G6) was applied to immunohistochemical detection of HLA-G5 and HLA-G6 molecular expression in different tissues.

[0049] The collected tissues were fixed in 10%-12% neutral formalin and embedded in paraffin. Tissue sections go through routine production processes such as baking, dewaxing, hydration, and antigen retrieval. An appropriate amount of 1% BSA was added dropwise on the tissue, covering the tissue and the edge of the tissue by 2 mm, and incubated at room temperature for 10 min to seal. Anti-HLA-G5 and HLA-G6 isoform antibody (9Y5G6) (1 mg / mL, diluted 1:500) was added dropwise, and kept overnight in a humid box at 4°C (16-20h). Rinse with TBS buffer, add secondary antibody dropwise (antibody goat-to-mouse ratio 1:300 diluted in TBS), and incubate in a 37°C incubator for 30min. Rinse with TBS buffer solution, add the chromogenic agent DAB working solution dropwise, and after the tissue color development is complete, rinse the slid...

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Abstract

The invention discloses an antibody (9Y5G6) against HLA-G isomer molecules HLA-G5 and HLA-G6 and its application. The antibody (9Y5G6) is produced by a hybridoma cell line 9Y5G6 with a deposit number of CCTCC NO: C202045 , using the antigenic peptide of the 229-319th amino acid sequence of the heavy chain of the HLA-G molecule as an immunogen, encoding the nucleic acid molecule and amino acid sequence of the 9Y5G6 antibody of the present invention, and the present invention also provides the 9Y5G6 antibody for use in HLA-G The use of isomeric molecules HLA-G5 and HLA-G6 immunohistochemistry and western blot detection.

Description

technical field [0001] The invention belongs to the field of biomedicine and relates to the following aspects of an anti-HLA-G isoform antibody (9Y5G6): the antigenic peptide of the 229-319th amino acid sequence of the heavy chain of an HLA-G molecule is used as an immunogen to prepare an anti-HLA-G isoform antibody Antibody (9Y5G6) of conformation HLA-G5 and HLA-G6; Nucleic acid molecule and amino acid sequence encoding the 9Y5G6 antibody of the present invention, and said antibody (9Y5G6) is used for HLA-G5 and HLA-G6 immunohistochemistry and immunohistochemistry blot detection, etc. Background technique [0002] Human leukocyte antigen-G (human leukocyte antigen-G, HLA-G) gene, with a full length of 6.0kb, is located at 6p21.3 on the distal side of the short arm of human chromosome 6. During protein translation, the first exon of HLA-G mRNA encodes the signal peptide, the second, third, and fourth exons encode the α1, α2, and α3 domains of the extracellular region, respe...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/28C12N15/13C12N5/20G01N33/577G01N33/574G01N33/569
CPCC07K16/2833G01N33/577G01N33/57492G01N33/57446G01N33/56983C07K2317/565C07K2317/567C07K2317/92G01N2333/70539
Inventor 颜卫华林爱芬章霞
Owner TAIZHOU ENZE MEDICAL CENT GROUP
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