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Binding protein binding to HBeAg and application of binding protein

A technology for binding proteins and binding domains, applied in the field of antibodies, which can solve the problems of difficult analysis, prone to false positives, and large differences between batches

Active Publication Date: 2021-06-18
DONGGUAN PENGZHI BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Traditional clinical diagnosis uses mouse-derived monoclonal antibodies, which are particularly affected by individual mice, with unstable production, large batch-to-batch differences, and difficult purification of mouse autoantibodies.
In addition, there are also some disadvantages in the use of mouse monoclonal antibodies in the detection process, especially when two mouse-derived monoclonal antibodies are used in the detection reagent of the double-antibody sandwich method, false positives are prone to occur, and the reasons for these false positives There are many, and it is difficult to analyze. For example, there may be HAMA effect in blood samples, and there may be some components that can bind to mice in some throat swabs or nasal swab samples
In addition, the existing anti-HBeAg antibodies cannot be well used in the detection of HBeAg due to their low activity and poor affinity.

Method used

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  • Binding protein binding to HBeAg and application of binding protein
  • Binding protein binding to HBeAg and application of binding protein
  • Binding protein binding to HBeAg and application of binding protein

Examples

Experimental program
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Effect test

Embodiment 1

[0118] In this embodiment, the resolved endonuclease is resolved, and the Prime STAR DNA polymerase is purchased from Takara. Magextractor-RNA extraction kit purchases from ToyoBO. BD smart TM Race CDNAMPLification Kit kit purchases from Takara. PMD-18T vectors were purchased from Takara. Plasmid extraction kits were purchased from Tiangen. Primer synthesis and gene sequencing were completed by Invitrogen.

[0119] This embodiment provides a method of preparing a recombinant antibody against HBeAg

[0120] 1 construction of recombinant plasmid

[0121] (1) primers

[0122] Amplification Heavy Chain and Light Chain 5'race Primers:

[0123]

[0124] (2) Gene cloning and sequencing of antibody variable region

[0125] RNA was extracted from hybridoma cell line with HBeAg monoclonal antibody, and the first chain cDNA synthesis was performed using Smarter II A OligonucleiTide and 5'-CDS primers in SmartTM RacecDNAMPLification Kit kits and kits. Chain cDNA product as a PCR amplificati...

Embodiment 2

[0147] Properties detection of antibodies

[0148] (1) Example 1 antibody and its mutant activity detection

[0149] Further analysis, the heavy chain variable region of the HBeAg monoclonal antibody (WT) of Example 1, as shown in SEQ ID NO: 13, wherein the amino acid sequence of each complementary determining region of the heavy chain is as follows:

[0150] CDR-VH1: G-Y-T (X1) -F-T-D (x2) -Y-N (x3) -M-N;

[0151] CDR-VH2: N-I (X1) -D-p-Y-F-G-S (X2) -S-D (x3) -Y-N-Q (X4) -k;

[0152] CDR-VH3: A (X1) -R-Y-L (x2) -T-W-D (x3) -Y-Y-a-M;

[0153] Its light chain variable region is shown in SEQ ID NO: 11, wherein the amino acid sequence of each complex determining area of ​​the light chain is as follows:

[0154] CDR-VL1: R-S-S-K (X1) -S-I (X2) -V-H-S-D (X3) -G-N-T-Y-L (X4) -e;

[0155] CDR-VL2: K-V-S-D (x1) -R-F (x2) -s;

[0156] CDR-VL3: F-Q (x1) -g-S-H-V (x2) -p-p.

[0157] Based on the HBeAg monoclonal antibody of Example 1, a displacement of antibody activity is performed in a com...

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Abstract

The invention discloses a binding protein binding to HBeAg and application of the binding protein, and relates to the technical field of antibodies. The binding protein specifically binding to HBeAg disclosed by the invention comprises an antigen binding structural domain, and the antigen binding structural domain comprises at least one of the following complementarity determining regions: a complementarity determining region CDR-VH1, a complementarity determining region CDR-VH2 and a complementarity determining region CDR-VH3. The binding protein specifically binding to the HBeAg disclosed by the invention can bind to the HBeAg, has higher binding activity and affinity, can be used for detecting the antigen level of the HBeAg and diagnosing HBV infection, and provides more protein choices for the detection of the HBeAg and the diagnosis of the HBV infection.

Description

Technical field [0001] The present invention relates to the field of antibody technology, specifically to binding proteins that bind HBeAg and their applications. Background technique [0002] Globally, there are approximately 2 billion people infected with hepatitis B virus (HBV), and it is the main cause of acute and chronic liver diseases. About 1 million people die from liver failure and liver failure caused by HBV infection every year. Cirrhosis and primary hepatocellular carcinoma. There are about 93 million people with chronic HBV infection in my country, of which about 20 million are chronic hepatitis B patients. Chronic hepatitis B still lacks effective control methods and complete solutions. The relatively lagging progress in research on the molecular mechanism of the interaction between HBV and liver cells and immune cells is one of the important reasons that seriously affects the development of new treatments and drugs. [0003] Hepatitis B e antigen (HBV e ant...

Claims

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Application Information

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IPC IPC(8): C07K16/08C12P21/02G01N33/576
CPCC07K16/082G01N33/5761C07K2317/56C07K2317/565C07K2317/92C07K2317/94G01N2333/02
Inventor 崔鹏何志强孟媛钟冬梅周俊覃婷
Owner DONGGUAN PENGZHI BIOTECH CO LTD
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