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Tuberculosis vaccine candidate component and vaccine containing tuberculosis vaccine candidate component

A technology for tuberculosis and Mycobacterium tuberculosis, applied in the fields of immunology and molecular biology, can solve the problems of unclear effective antigenic components of genome and proteome, basic research needs to be strengthened, and no gene annotation.

Active Publication Date: 2017-10-24
INST OF PATHOGEN BIOLOGY CHINESE ACADEMY OF MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The microcart, obtained after long-term screening, is the only effective therapeutic vaccine against tuberculosis so far. Although its application prospect is very promising, basic research on this vaccine strain needs to be strengthened, for example, its genome, proteome, and effective antigenic components etc. are not clear; in addition, its performance characteristics are very different from traditional BCG, and the genetic background of BCG has no reference; the mechanism of microcart to play an immune role to achieve therapeutic purposes is also unclear
[0005] Genomic and proteomic research on Mycobacterium vaccae has become one of the research hotspots. The draft genome (Scaffolds or contigs) of M. vaccae ATCC 25954 has been submitted to the NCBI database, but there is no complete genome fine map, nor There is no accurate gene annotation, which brings great obstacles to the follow-up research of such vaccines[7]

Method used

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  • Tuberculosis vaccine candidate component and vaccine containing tuberculosis vaccine candidate component
  • Tuberculosis vaccine candidate component and vaccine containing tuberculosis vaccine candidate component
  • Tuberculosis vaccine candidate component and vaccine containing tuberculosis vaccine candidate component

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1: Preparation and Identification of Protein MYVA_1927

[0044] The preparation of the protein MYVA_1927 (shown in SEQ ID NO:1) can be obtained by obtaining its coding base sequence (shown in SEQ ID NO:2), cloning it into an expression vector, and purifying it through protein expression. The coding base sequence can be prepared through synthetic chemical synthesis, and can also be cloned from Mycobacterium vaccae strain ATCC95051.

[0045] Specifically, the following methods can be used.

[0046] 1. Bacterial culture

[0047] Inoculate the frozen strain (ATCC 95051) into 200 ml of 7H9 liquid medium (10% OADC nutritional supplement, produced by BD Company, USA), cultivate overnight at 30°C, 100 rpm, subculture an appropriate amount according to 1:100 times, when the OD value When it reaches 0.8-1.0, place it on ice for 1.5 hours, centrifuge at 3000g for 10 minutes, and collect the strains.

[0048] A part of the collected strains was used for the genome extr...

Embodiment 2

[0095] Example 2: Protein MYVA_1927 specifically causes antibody production in tuberculosis patients

[0096] 1. Experimental materials

[0097] The strain collected in step 1 in Example 1.

[0098] Tuberculosis patient plasma: from 200 tuberculosis patients. Collected after informed consent of subjects. The determination of the patient meets any one of the following conditions: sputum smear or bacterial culture is positive; PCR amplification is positive (the target gene of PCR amplification is the conserved sequence fragment IS6110 of Mycobacterium tuberculosis, the copy number is 2x10 5 positive); chest X-ray or CT support is a tuberculosis patient; clinical ESAT-6 positive.

[0099] Healthy human plasma: from 50 healthy people. Collected after informed consent of subjects. The determination of healthy people meets all the following conditions: no clinical symptoms and chest X-ray confirmed healthy people, and ESAT-6 is negative.

[0100] 2. Experimental method

[0...

Embodiment 3

[0149] Embodiment 3: cellular immunity test

[0150] ELISPOT analysis enables the visualization of single-cell secreted products. These assays are exceptionally sensitive because it is captured directly around the secreting cell and before the surface is diluted, or captured by receptors on adjacent cells, or degraded. Autoimmune testing, organ transplantation, vaccine and drug development and testing, T cell function research, tumor research, infectious disease research and testing, virus research, etc. The specific principle is: cells are stimulated by antigens to produce cytokines, and the cytokine antigens are captured by specific monoclonal antibodies. After cell dissociation, the captured cytokines are bound to a biotin-labeled secondary antibody, followed by alkaline phosphatase-labeled avidin. After incubation with BCIP / NBT substrate, "purple" spots appeared on the PVDF plate, indicating that the cells had produced cytokines, and the results were obtained after ana...

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Abstract

The invention belongs to the fields of immunology and molecular biology and relates to a tuberculosis vaccine candidate component and a vaccine containing the tuberculosis vaccine candidate component. The invention further relates to a molecular marker diagnosing tuberculosis. Specifically, the invention relates to application of a protein MYVA_1927 in preparation of drugs for treating and / or preventing tuberculosis or drugs for inhibiting mycobacterium tuberculosis, or application in preparation of drugs for diagnosing tuberculosis. The invention further relates to a vaccine containing the protein MYVA_1927. The protein MYVA_1927 has excellent coincidence rate and response intensity and has potential of being applied to preparing the tuberculosis vaccine. In addition, the protein MYVA_1927 can further serve as an immunologic diagnosis molecular marker of tuberculosis (such as pulmonary tuberculosis).

Description

technical field [0001] The invention belongs to the field of immunology and molecular biology, and relates to a candidate component of a tuberculosis vaccine and a vaccine containing the component. The invention also relates to a molecular marker for diagnosing tuberculosis. Background technique [0002] Tuberculosis is one of the major infectious diseases that endanger human health, and it is the main killer of adult infectious diseases in the world today, which has posed a severe challenge to international public health [1]. my country is one of the 22 countries with a high burden of tuberculosis in the world. The epidemic situation shows "three highs and one low", that is, high prevalence rate, high mortality rate, high drug resistance rate and low annual decline rate. And with the emergence of AIDS co-infection, drug-resistant tuberculosis and the increase of floating population, the situation of tuberculosis epidemic is becoming more and more serious, and the situation...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/16A61K39/04A61K39/39A61K45/06A61P31/06A61P11/00G01N33/569G01N33/68G01N33/561G01N33/94C12Q1/68
CPCA61K38/164A61K39/04A61K39/39A61K45/06A61K2039/575C12Q1/6883C12Q2600/158G01N33/561G01N33/5695G01N33/6848G01N33/94
Inventor 金奇郑建华张笑冰陈丽宏杨剑刘立国
Owner INST OF PATHOGEN BIOLOGY CHINESE ACADEMY OF MEDICAL SCI
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