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Hepatitis B fusion protein multiva lentvaccine, preparation method and uses thereof

A fusion protein, hepatitis B technology, applied in the field of multivalent vaccine preparation, can solve the problems of pollution, difficult quality inspection standards, etc.

Inactive Publication Date: 2008-06-25
SHANGHAI NAT ENG RES CENT OF ANTIBODY MEDICINE +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, since the fully human anti-HBsAg polyclonal antibody in IC is mainly derived from anti-HBsAb positive human serum (Liu SA, Xu DZ, Zhang JP, et al.Immune response for phase I clinical trial of a hepatitisB immunogenic complex therapeutic vaccine, YIC.Zhonghua Gan Zang BingZa Zhi, 2006; 14(2):89-92), with the increasing demand for vaccines, the source of hepatitis B surface antibody positive sera may become a bottleneck for large-scale production, and the current use of The immune complex vaccine prepared from the patient's serum-derived antibody has potential contamination, and it is difficult to have a unified quality inspection standard for the immune complex vaccine prepared from antibodies from different sera

Method used

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  • Hepatitis B fusion protein multiva lentvaccine, preparation method and uses thereof
  • Hepatitis B fusion protein multiva lentvaccine, preparation method and uses thereof
  • Hepatitis B fusion protein multiva lentvaccine, preparation method and uses thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Example 1. PCR of N-terminal fusion Flt3 ligand extracellular segment gene

[0046] Referring to the information and sequence of human Flt3 ligand provided by GENEBANK, synthetic primers: FL primer sense (primer 1): GCG ATGACAGTGCTGGCGCCA (Hind III) and FL antisense (primer 2): CGGGGCTGTCGGGGCTGT (Shaded part is complementary to Hinge 5'; underlined part is complementary to FL 3'). Human T lymphocytes were isolated, RNA was extracted with TRISOL kit (INVITROGEN), and a 546bp DNA fragment was obtained by RT-PCR (PROMEGA). After the fragments were recovered by a gel recovery kit (Shanghai Sangong), they were used for later use. The nucleotide sequence of the Flt3 ligand extracellular segment gene is shown in SEQ ID NO.1.

Embodiment 2

[0047] Example 2. PCR of human antibody heavy chain constant region Hinge+CH2+CH3 gene

[0048] Use lymphocyte separation medium to separate healthy human lymphocytes, use Trizol reagent (Invitrogen company product) to extract total RNA, and design primers for the Hinge+CH2+CH3 gene of the constant region of the heavy chain of the human antibody: Hinge+CH2+CH3 sense (primer 3) : GAG CCC AAA TCT TGT GAC (this sequence is consistent with Hinge5') and Hinge+CH2+CH3 antisense (primer 4): GCG TTT ACC CGG AGA CAGGGA (Nhe I, this sequence is complementary to CH3 3'), amplifies the antibody heavy chain constant region gene 696bp. The PCR reaction adopts hot start, and the reaction conditions are: 94°C for minutes; 94°C for 45 seconds, 60°C for 45 seconds, 72°C for 1 minute and 10 seconds, 30 cycles; 72°C for 10 minutes. The PCR products were purified and recovered by agarose gel electrophoresis. The heavy chain CH2-CH3 gene sequence of human IgG1 with hinge region is shown in SEQ I...

Embodiment 3

[0049] Example 3. Cloning of FL-Hinge-CH2-CH3

[0050] FL and Hinge+CH2+CH3 were used as templates, FL sense (primer 1) and Hinge+CH2+CH3 antisense (primer 4), amplified by overlap PCR to obtain FL+Hinge+CH2+CH3 (1242bp), and cloned into In the pGEM-T vector, it was confirmed that the correct clone was obtained after sequencing verification. The correct clone in this example was designated pGEM-T / FL-Hinge-CH2-CH3.

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Abstract

This invention discloses a multivalence hepatitis B fusion protein targeting adjuvant vaccine, a preparation method and an application thereof. Particularly, the invention discloses a quadrivalence hepatitis B fusion protein targeting adjuvant vaccine (FL-CH-HBsAg-HbsAg)2, an octivalence hepatitis B fusion protein targeting adjuvant vaccine (HBsAg-HBsAg-CH-FL, HBsAg-HBsAg-CL)2, a relevant preparation method, and an application in preparation of preventative pharmaceuticals, and in treatment and / or prevention of hepatitis B.

Description

technical field [0001] The invention belongs to the field of bioengineering, and more specifically, the invention discloses a multivalent vaccine, its preparation method and application. Background technique [0002] Hepatitis B virus is the most common infectious disease in the world. At present, about 400 million people in the world are carriers of chronic hepatitis B virus, while China has about 100 million carriers, ranking first in the world. my country is also a country with a high incidence of hepatitis B virus infection. There are about 30 million chronic hepatitis B patients, and the hepatitis B infection rate in the population is as high as 57%. Among them, 1 / 4 of the hepatitis B virus carriers may develop into hepatocellular carcinoma, and 80% of the liver cells Cancer is due to hepatitis B virus (hepatitis B virus hereinafter referred to as HBV) infection. Since the early 1980s, the vaccination of hepatitis B vaccine has achieved great success in preventing and ...

Claims

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Application Information

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IPC IPC(8): A61K39/395A61K39/39C12N15/62A61P1/16A61P31/14A61P31/20A61K39/29
Inventor 郭亚军戴建新李博华侯盛钱卫珠王皓
Owner SHANGHAI NAT ENG RES CENT OF ANTIBODY MEDICINE
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