Mycobacterium tuberculosis specificity CTL epitope peptides and application thereof

A technology of Mycobacterium tuberculosis and epitope peptides, which is applied in the fields of molecular biology and immunology, can solve the problems of unclear characteristics of tuberculosis vaccines, no MHC typing of peptides, affinity and stability to be identified, etc.

Inactive Publication Date: 2017-05-31
THE 309TH HOSPITAL OF CHINESE PEOPLES LIBERATION ARMY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] Published on June 27, 2012, the patent application number is CN201110362902.6, and the Chinese invention patent titled "Epitope Polypeptides Usable for Detecting Mycobacterium Tuberculosis Infection and Its Application" discloses several Mycobacterium tuberculosis-specific Sexual antigenic epitope polypeptides, especially the antigenic epitope polypeptides of antigen Rv3615c, are 8-11 continuous polypeptide fragments. Although the antigenic polypeptides can generate T cell responses in tuberculosis patients, these polypeptides have neither MHC typing nor For CTL epitope identification, the characteristics of using it as a tuberculosis vaccine are not clear, its affinity and stability need to be identified, and the in vitro immune activity needs to be improved

Method used

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  • Mycobacterium tuberculosis specificity CTL epitope peptides and application thereof
  • Mycobacterium tuberculosis specificity CTL epitope peptides and application thereof
  • Mycobacterium tuberculosis specificity CTL epitope peptides and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1: Preparation of HLA-A2-restricted CTL epitope polypeptide of Mycobacterium tuberculosis-specific protein Rv2629

[0036] 1. Prediction of HLA-A2-restricted CTL epitopes of Mycobacterium tuberculosis-specific protein Rv2629 using bioinformatics methods

[0037] Mycobacterium tuberculosis specific protein Rv2629, its amino acid composition is shown in SEQ ID NO.13.

[0038] SEQ ID NO.13:

[0039]MRSERLRWLVAAEGPFASVYFDDSHDTLDAVERREATWRDVRKHLESRDAKQELIDSLEEAVRDSRPAVGQRGRALIATGEQVLVNEHLIGPPPATVIRLSDYPYVVPLIDLEMRRPTYVFAAVDHTGADVKLYQGATISSTKIDGVGYPVHKPVTAGWNGYGDFQHTTEEAIRMNCRAVADHLTRLVDAADPEVVFVSGEVRSRTDLLSTLPQRVAVRVSQLHAGPRKSALDEEEIWDLTSAEFTRRRYAEITNVAQQFEAEIGRGSGLAAQGLAEVCAALRDGDVDTLIVGELGEATVVTGKARTTVARDADMLSELGEPVDRVARADEALPFAAIAVGAALVRDDNRIAPLDGVGALLRYAATNRLGSHRS

[0040] 1. SYFPEITHI supermotif method for remote prediction of CTL epitopes:

[0041] Using SYFPEITHI online software (Ver.1.0) to predict the CTL epitope consisting of 9 amino acid residues;

[00...

experiment example 1

[0084] Experimental example 1: In vitro immune activity detection of epitope polypeptides

[0085] 1. Materials and methods

[0086] 1. Experimental materials

[0087] 1.1 Experimental cell lines

[0088] The T2 cell line was donated by the Department of Immunology, Third Military Medical University, and cultured in RPMI1640 medium containing 10% fetal bovine serum in a 37°C, 5% carbon dioxide incubator.

[0089] 1.2 Peripheral blood samples

[0090] Peripheral blood samples were provided by the Army Tuberculosis Research Institute and were divided into normal control group, LTBI group and tuberculosis group. The HLA phenotype analysis of peripheral blood was detected by flow cytometry in the organ transplantation laboratory, all of which were positive for HLA-A2.

[0091] 1.3 Main reagents

[0092] RPMI1640 medium: product of Gibco, USA;

[0093] IMDM medium: product of Gibco, USA;

[0094] Fetal bovine serum: product of Gibco, USA;

[0095] Glutamine: product of Sigma...

experiment example 2

[0164] Experimental example 2: Lactate dehydrogenase method (Lactate dehydrogenase, LDH) detection of specific CTL cytotoxic killing experiment

[0165] 1. Materials and methods

[0166] 1. Experimental materials

[0167] 1.1 Experimental cell lines

[0168] The T2 cell line was donated by the Department of Immunology, Third Military Medical University of the Chinese People's Liberation Army, and cultured in RPMI1640 medium containing 10% fetal bovine serum in a 37°C, 5% carbon dioxide incubator.

[0169] 1.2 Peripheral blood samples

[0170] Peripheral blood samples were provided by the Army Tuberculosis Research Institute and were divided into normal control group, LTBI group and tuberculosis group. The HLA phenotype analysis of peripheral blood was detected by flow cytometry in the organ transplantation laboratory, all of which were positive for HLA-A2.

[0171] 1.3 Main reagents

[0172] RPMI1640 medium: product of Gibco, USA;

[0173] IMDM medium: product of Gibco, U...

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Abstract

The invention relates to mycobacterium tuberculosis specificity CTL epitope peptides and application thereof and belongs to the field of molecular biology and immunology. Through prediction of biological informatics software, selection of HLA-A2 limitation CTL epitope, modification of partial epitope peptides and synthesis and activity comparison and screening of epitope peptides, the obtained four 9-peptides of Rv2629-p190-2L, Rv2629-p190-1Y2L, Rv2629-p274 and / or Rv2629-p315 of the HLA-A2 limitation CTL epitope of mycobacterium tuberculosis protein Rv2629 have higher affinity and higher stability; the IFN-gamma levels for inducing PBMCs generation of a tuberculosis patient are obviously higher than that of a normal control group and a tuberculosis dormant infection group; the CTL ability for stimulating the PBMCs generation of the tuberculosis patient to secrete IFN-gamma is obviously higher than that of the normal control group and the tuberculosis dormant infection group; all the four 9-peptides can induce to generate CTL with killing activity to target cells, and the killing activity can be correspondingly improved along with increasing of effect and target ratio; CTL generated by stimulating modified peptides of Rv2629-p190-2L and Rv2629-p190-1Y2L can identify mother peptide Rv2629-P190. The mycobacterium tuberculosis specificity CTL epitope peptides can serve as a tuberculosis cell-mediated immunity inducer and can be a novel candidate epitope of antituberculous epitope vaccine or an immunity diagnosis reagent.

Description

technical field [0001] The present invention relates to an epitope polypeptide and its application, in particular to a mycobacterium tuberculosis protein (Rv2629) specific cytotoxic T lymphocyte (Cytotoxic T lymphocyte, CTL) epitope polypeptide and its application. These epitope polypeptides can The method can cause specific cellular immune response of tuberculosis patients, has application value in the research and development of tuberculosis immunodiagnostic reagents and epitope vaccines, and belongs to the fields of molecular biology and immunology. Background technique [0002] Tuberculosis is an important public health problem and one of the major infectious diseases threatening human health worldwide, with high morbidity and high mortality. The prevalence of AIDS, the flow of patients with bacterium-expelling tuberculosis, and the poverty of some people have led to an upward trend in the incidence of tuberculosis. In particular, problems such as drug resistance of Myco...

Claims

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Application Information

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IPC IPC(8): C07K7/06G01N33/68G01N33/569A61K39/04A61P31/06
CPCA61K39/04C07K7/06G01N33/5695G01N33/68G01N2333/35
Inventor 吴雪琼白雪娟王东方林明贵刘银萍梁艳阳幼荣
Owner THE 309TH HOSPITAL OF CHINESE PEOPLES LIBERATION ARMY
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