Anti-hbs monoclonal antibodies

A monoclonal antibody and antibody technology, applied in the direction of anti-animal/human immunoglobulin, cells modified by introducing foreign genetic material, biochemical equipment and methods, etc., can solve the problems of infection transmission, affecting HBV hosts, etc.

Active Publication Date: 2007-01-17
SYSMEX CORP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

If HBsAg containing mutations (mutated HBsAg) cannot be detected, not only the host of HBV ...

Method used

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Examples

Experimental program
Comparison scheme
Effect test

experiment example 1

[0090] (1) Preparation of monoclonal antibodies

[0091] 1-1. Immunization of mice

[0092] 100 μl of FCA was added to 100 μl of PBS containing 50 μg of inactivated adr-type HBsAg (manufactured by TRINA) purified from human serum, and mixed and emulsified with a Voltex vortex to prepare 200 μl of an FCA-mixed HBsAg solution. In addition, 200 µl of an FIA-mixed HBsAg solution was prepared in the same manner except that FIA was used instead of FCA.

[0093] Primary immunization was performed by injecting 200 μl of FCA mixed with HBsAg solution into the abdominal cavity of 8-week-old female BALB / c mice. After the initial immunization, 200 μl of FIA mixed HBsAg solution was used for five booster immunizations every two weeks. Four days after the last booster immunization, splenocytes were isolated, and the cells were fused with P3X63-Ag8·653 mouse myeloma cells using the PEG method to prepare hybridomas.

[0094] 1-2. Culture of hybridoma

[0095] Suspend the hybridomas in HT ...

experiment example 2

[0108] Using DNA of wild-type HBsAg as a template, DNA encoding the amino acid sequence from position 1 to position 196 of wild-type HBsAg (subtype: adr) was prepared by PCR method and inserted into the expression vector pcDNA3.1(+) for eukaryotic cells , A plasmid for expressing wild-type HBsAg was prepared. Introduce the plasmid into COS7 cells, in 5% CO 2 Incubate in an incubator for 24 hours. The COS7 cells can express HBsAg with amino acid deletion below 197 (hereinafter referred to as C-terminal deletion HBsAg).

[0109] Whether or not the 149 antibody and the 1053 antibody react with the C-terminal-deleted HBsAg was determined in the same manner as in Experimental Example 1. The judgment results are shown in Table 3 below.

[0110] Antibody 149

[0111] From Table 3, it can be confirmed that the 149 antibody and the 1053 antibody can recognize the C-terminal deletion HBsAg. From this, it was found that the epitopes of the 149 antibody and the 1053 antibo...

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PUM

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Abstract

An anti-HBs monoclonal antibody is described herein. This antibody can bind to the following: a wild type HBsAg; at least one mutant HBsAg selected from the group consisting of a first mutant HBsAg and a second mutant HBsAg; and at least one mutant HBsAg selected from the group consisting of a third mutant HBsAg and a fourth mutant HBsAg. The first mutant HBsAg has a mutation at position 120. The second mutant HBsAg has a mutation at position 141. The third mutant HBsAg has a substitution to a lysine at position 118. The fourth mutant HBsAg has only one mutation at position 144 and an amino acid at the position 144 is substituted by a glutamic acid.

Description

technical field [0001] The present invention relates to monoclonal antibodies specifically binding to HBsAg. Specifically, it relates to monoclonal antibodies that bind to wild-type HBsAg as well as mutant HBsAg. Background technique [0002] Hepatitis B virus (HBV) is a double-stranded DNA virus of the family Hepadnaviridae having an outer sheath of a lipid bilayer membrane, a core shell as a protein, a DNA polymerase, and genomic DNA. S protein, M protein and L protein are combined in the outer sheath, and the surface antigen (HBsAg) located on the S protein is related to virus infection. HBsAg is a polypeptide composed of 226 amino acids, and there are four subtypes of adw, ayw, adr and ayr in HBsAg. adw, the 122nd and 160th amino acids of HBsAg are both lysine. ayw, the amino acid at position 122 of HBsAg is arginine, and the amino acid at position 160 is lysine. adr, the amino acid at position 122 is lysine, and the amino acid at position 160 is arginine. ayr, the ...

Claims

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Application Information

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IPC IPC(8): C07K16/18C12N5/12C12N5/18
CPCG01N33/576G01N2333/02C07K16/082
Inventor 武田和彦梶田忠宏朝枝步
Owner SYSMEX CORP
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