Mycobacterium tuberculosis antigen protein Rv1798 and application of T cell epitope peptide of Mycobacterium tuberculosis antigen protein Rv1798
A technology of Mycobacterium tuberculosis, rv1798, applied in the fields of molecular biology and immunology, can solve the problems of poor sensitivity, high price, poor specificity of tuberculosis, etc., and achieve the effect of facilitating quality control, improving sensitivity and reducing false positives
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Embodiment 1
[0043] Example 1 Cloning of Mycobacterium tuberculosis antigen gene Rv1798 and expression and purification of protein
[0044] Rv1798 (GI: 15608935) is a conserved membrane protein encoded by the Mycobacterium tuberculosis H37Rv genome, containing 610 amino acids, and its amino acid sequence is shown in SEQ ID NO:3. According to its coding gene sequence, primers were designed, and the antigenic protein Rv1798 was expressed and purified using a prokaryotic expression system (such as Escherichia coli).
Embodiment 2
[0045] Example 2 Synthesis of Mycobacterium tuberculosis antigenic protein Rv1798T 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 Rv1798 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 antigenic protein Rv1798T cell epitope peptide provided in this example is selected from P308 and P309, and its amino acid sequences are respectively shown in SEQ ID NO: 1-2.
Embodiment 3
[0048] The preparation of embodiment 3 tuberculosis T-SPOT detection kit
[0049] The basic composition of the test kit is as follows:
[0050] ①The protein antigen Rv1798 prepared in Example 1 and / or the epitope peptide synthesized in Example 2
[0051] Antigen: the epitope peptide is selected from at least one of P308 and P309.
[0052] ② Primary antibody: mouse IgG monoclonal antibody against human or animal IFN-γ.
[0053] Enzyme-labeled reagent: another mouse IgG monoclonal antibody labeled with horseradish peroxidase against different epitopes of human or animal IFN-γ.
[0054] ③Standard product:
[0055] 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.
[0056] ④ Other reagents and consumables required for T-SPOT detection.
[0057] The primary antibody was immobili...
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Abstract
Description
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Application Information
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