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Mycobacterium tuberculosis antibody binding peptide and application thereof

A technology of Mycobacterium tuberculosis and peptide binding, applied in the field of medicine

Active Publication Date: 2011-07-13
SHANGHAI PULMONARY HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the application of phage display random peptide library technology to directly screen the specific antibody-binding peptides of tuberculosis patients has not been reported at home or abroad (according to VIP, PubMed and other Chinese and English database searches)

Method used

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  • Mycobacterium tuberculosis antibody binding peptide and application thereof
  • Mycobacterium tuberculosis antibody binding peptide and application thereof
  • Mycobacterium tuberculosis antibody binding peptide and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Example 1: Screening and sequence analysis of specific antibody-binding peptides in tuberculosis patient serum.

[0018] Experimental Materials.

[0019] 1. Phage Random Peptide Library Kit

[0020] Phage Surface Capsid Protein III Display Linear 7-peptide Library Kit (Ph.D.-7TMPhage Display Peptide Library Kit) was purchased from New England Biolabs, USA. The titer of the peptide library was 2 × 10 11 pfu / ml. Recipient bacteria E. coli The genotype of ER2738 was F’ lacIq Δ(lacZ)M15 proA+B+ zzf::Tn 10 (TetR) / fhuA2 supE thi Δ(lac-proAB)Δ(hsdMS-mcrB)5 (rk – m k – McrBC –). DNA automatic sequencing primer (-96III sequencing primer): 5'-CCCTCATAGTTAGCGTA ACG -3' (SEQ ID NO.1).

[0021] 2. Serum from tuberculosis patients and healthy people

[0022] The sera of 47 tuberculosis patients were collected from tuberculosis outpatients or hospitalized tuberculosis patients of Shanghai Pulmonary Hospital. All had a history of BCG vaccination. All candidates were excluded ...

Embodiment 2

[0082] Example 2: Detection of positive single phages on serum samples.

[0083] Experimental Materials.

[0084] Serum samples: Serum samples from 47 patients with active pulmonary tuberculosis (sputum samples were all positive for Mycobacterium tuberculosis culture) and 37 healthy people with a history of BCG vaccination were all preserved in the serum sample bank of Shanghai Pulmonary Hospital.

[0085] Main reagents: HRP-labeled goat anti-human IgG antibody was purchased from Sigma, USA

[0086] Preparation of related reagents:

[0087] Coating solution: 17.5 mmol / L Na2CO3, 32.5 mmol / L NaHCO3, 15 mmol / LMgCl2 6H2O, pH 9.6

[0088] PBS buffer: 0.02mol / L Na2HPO4, 0.02mol / L NaH2PO4.

[0089] Enzyme working solution: 3% BSA, PBS buffer

[0090] equipment:

[0091] ROTINA38 desktop refrigerated high-speed centrifuge (Hettich, Germany); Thermo MK3 microplate reader (Thermo, USA); HZQ-C air bath oscillator (Harbin Donglian Electronic Technology Development Co., Ltd.); fully a...

Embodiment 3

[0107] Example 3: Application of antibody-binding peptides in serological detection of tuberculosis.

[0108] Experimental Materials.

[0109] Serum samples: Serum samples from 47 patients with active pulmonary tuberculosis (sputum samples were all positive for Mycobacterium tuberculosis culture) and 37 healthy people with a history of BCG vaccination were all preserved in the serum sample bank of Shanghai Pulmonary Hospital.

[0110] 3.1 Polypeptide synthesis: According to the serological test results of the single phage, select the polypeptide displayed by the single phage T18 with better test results (named TP18), and the peptide sequence is LPPPPPFS, and entrust the company to synthesize in vitro.

[0111] 3.2 Polypeptide dissolution: Take a part of the dry peptide powder, weigh it accurately, dissolve it with ultrapure water to a 10 mg / ml solution, and store it at 4°C.

[0112] 3.3 Judgment of coating concentration: Dilute the peptide concentration to 0.5μg / ml~32μg / ml (0...

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Abstract

The invention belongs to the technical field of medicaments and in particular relates to mycobacterium tuberculosis antibody binding peptide and application thereof. The general formula of an amino acid sequence is X1X2X3PPFS, wherein P represents proline; F represents phenylalanine; S represents serine; X1 to X3 represent any amino acid; preferably, X1 is L or F, X2 is P or H, and X3 is P or F; and L represents leucine and H represents histidine. The tuberculosis patient serum specific antibody-bound polypeptide can be used as a detection reagent in tuberculosis serological detection.

Description

technical field [0001] The invention belongs to the technical field of medicine, and in particular relates to a polypeptide capable of binding to serum-specific antibodies of tuberculosis patients and the application of the polypeptide in serological detection of tuberculosis. Background technique [0002] Tuberculosis is an infectious disease controlled by WHO, and it is still a public health problem that seriously endangers people's health. According to relevant statistics, about 1 / 3 of the world's population is infected with tuberculosis, and there are about 20 million tuberculosis patients, with 8 to 10 million new cases every year, and about 2 million people die of tuberculosis every year. The total number of deaths. The tuberculosis epidemic in my country is extremely severe. There are currently 500 million people infected with tuberculosis, accounting for 1 / 4 of the world's total. Every year, 130,000 people die from tuberculosis, which is twice the total number of de...

Claims

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Application Information

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IPC IPC(8): C07K7/06G01N33/53G01N33/569
Inventor 杨华胡忠义刘忠华秦莲花杨焕森
Owner SHANGHAI PULMONARY HOSPITAL
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