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Use of mycobacterium tuberculosis antigen protein Rv2941 and its T cell epitope peptide

A technology of mycobacterium tuberculosis and antigenic protein, which is applied in the fields of molecular biology and immunology, can solve the problems that have not yet met the needs of tuberculosis diagnosis, and achieve the effects of quality control, false positive reduction and high purity

Active Publication Date: 2017-02-15
ICDC CHINA CDC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In vitro gamma interferon detection based on T cells is used for auxiliary diagnosis of tuberculosis. This detection method can not only screen out active tuberculosis patients, but also detect latent patients, so as to better prevent and control latent tuberculosis At present, there are commercial IGRA detection kits that use the whole protein or polypeptide of the tuberculosis-specific antigens ESAT-6 and CFP-10 encoded by the RD1 region of the Mycobacterium tuberculosis genome as stimuli, such as QuantiFERON-TB Gold test and T -SPOT, both exhibit high sensitivity and specificity, but have not yet met the diagnostic requirements of tuberculosis

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  • Use of mycobacterium tuberculosis antigen protein Rv2941 and its T cell epitope peptide
  • Use of mycobacterium tuberculosis antigen protein Rv2941 and its T cell epitope peptide
  • Use of mycobacterium tuberculosis antigen protein Rv2941 and its T cell epitope peptide

Examples

Experimental program
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Effect test

Embodiment 1

[0043] Example 1 Cloning of Mycobacterium tuberculosis antigen gene Rv2941 and protein expression and purification

[0044] Rv2941 (GI: 15610078) is a conserved membrane protein encoded by the Mycobacterium tuberculosis H37Rv genome, containing 580 amino acids, and its amino acid sequence is shown in SEQ ID NO:5. According to its coding gene sequence, primers are designed, and the antigenic protein Rv2941 is obtained by expressing and purifying using a prokaryotic expression system (such as Escherichia coli).

Embodiment 2

[0045] Example 2 Synthesis of Mycobacterium tuberculosis antigenic protein Rv2941 T cell epitope peptide

[0046] Based on the T cell IFN-γ release technology, the bioinformatics software TE predict and IEDB were used to predict the T cell antigen epitope on the Rv2941 coding gene, and the epitope peptide was synthesized by the solid-state synthesis method, and then the tuberculosis patients, Specific T lymphocytes in patients with other lung diseases and healthy people were detected, so as to evaluate the sensitivity and specificity of the antigen for tuberculosis detection.

[0047] The Mycobacterium tuberculosis antigen protein Rv2941 T cell epitope peptide provided in this example is selected from P327, P330, P332, and P334, and its amino acid sequences are shown in SEQ ID NO: 1-4 respectively.

Embodiment 3

[0048] Example 3 Preparation of Tuberculosis T-SPOT Detection Kit

[0049] The basic composition of the test kit is as follows:

[0050] ① The protein antigen Rv2941 prepared in Example 1 and / or the epitope peptide antigen synthesized in Example 2: the epitope peptide is selected from at least one of the amino acid sequences of SEQ ID NO: 1-4.

[0051] ② Primary antibody: mouse IgG monoclonal antibody against human or animal IFN-γ.

[0052] Enzyme-labeled reagent: another mouse IgG monoclonal antibody labeled with horseradish peroxidase against different epitopes of human or animal IFN-γ.

[0053] ③Standard product:

[0054] Culture plate: 96-well microwell reaction plate containing PVDF membrane or nitrocellulose membrane, positive control wells contain tuberculosis non-specific stimulating antigen (such as PHA, etc.), negative control wells contain PBS or base solution.

[0055] ④ Other reagents and consumables required for T-SPOT detection.

[0056] The primary antibody...

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Abstract

The invention relates to a use of mycobacterium tuberculosis antigen protein Rv2941 and its T cell epitope peptide in preparation of a tuberculosis detection agent, vaccine and drug. The antigen protein Rv2941 and its T cell epitope peptide have amino acid sequences respectively shown in the formulas of SEQ ID NO: 1 to 5. The mycobacterium tuberculosis Rv2941 protein antigen and its T cell epitope peptide as irritants are used for mycobacterium tuberculosis infection-caused specific T cell and B cell immunoreaction. Compared with the existing complete antigen, the mycobacterium tuberculosis antigen protein Rv2941 and its T cell epitope peptide can reduce a false positive result caused by antigen impurities. A detection reagent prepared from the Rv2941 protein antigen and its epitope peptide can be widely used in the related area of tuberculosis auxiliary diagnosis and epidemiological surveillance. A tuberculosis vaccine and an anti-tuberculosis drug prepared from the Rv2941 protein antigen and its epitope peptide can be used for tuberculosis prevention and treatment.

Description

technical field [0001] The invention relates to the fields of molecular biology and immunology, in particular to the application of mycobacterium tuberculosis antigen protein Rv2941 and its T cell epitope peptide in the preparation of tuberculosis detection reagents, vaccines and medicines. Background technique [0002] Tuberculosis is a chronic infectious disease caused by Mycobacterium tuberculosis. Survey results show that one-third of the world's population is latently infected, and 5% to 10% may develop active tuberculosis in future life. Since the World Health Organization declared tuberculosis a global crisis in 1993, the morbidity and mortality of tuberculosis have remained high. According to the WHO report, there are about 8 million new tuberculosis patients every year, and about 2 to 3 million people die of tuberculosis every year. my country ranks second among the 22 countries with a high burden of tuberculosis. The results of the fifth national epidemiological sa...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/569A61K39/04A61K39/40A61P31/06C07K14/35
CPCA61K39/00A61K39/04A61K2039/53C07K14/35G01N33/5695G01N33/68
Inventor 万康林刘海灿王雪枝李马超
Owner ICDC CHINA CDC
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