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Antibody combined with tubercle bacillus bacteriophage tail protein as well as preparation method and application thereof

A Mycobacterium tuberculosis and bacteriophage technology, applied in the preparation of antibodies, combined with the antibody field of Mycobacterium tuberculosis bacteriophage tail protein, can solve the problems of indistinguishable, indistinguishable cases, difficult to meet the sensitivity and specificity requirements, etc., to shorten the time and improve the sensitivity Effect

Inactive Publication Date: 2013-10-23
ABBOTT DIAGNOSTICS (SHANGHAI) CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the common problem of the above methods is that the sensitivity and specificity are difficult to meet the requirements, and they cannot distinguish the cured cases of previous infection; some products even cannot distinguish the cases of Bacillus Calmette-Guerin (BCG) injection

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1, Preparation of an antibody that recognizes the tail protein of Mycobacterium tuberculosis phage D29

[0033] 1. Synthesis of tail protein fragments of Mycobacterium tuberculosis phage D29

[0034] The nucleotide sequence of the tail protein fragment of Mycobacterium tuberculosis D29 is shown in SEQ ID NO:1, and the amino acid sequence of the tail protein fragment of Mycobacterium tuberculosis D29 is shown in SEQ ID NO:2.

[0035] The nucleic acid containing the sequence of SEQ ID NO: 1 was inserted into the pcDNA3 vector (purchased from Invitrogen) by molecular cloning, the correct clone was selected for amplification after sequencing and identification, the DNA was extracted and transformed into Escherichia coli, and the D29 tail protein fragment was expressed and purified.

[0036] 2. Preparation of antibodies that recognize D29 tail protein fragments

[0037] a) Animal immunization

[0038] Mix the previously purified D29 tail protein fragment with an eq...

Embodiment 2

[0045] Embodiment 2, the amplification and detection of D29 bacteriophage in Mycobacterium tuberculosis and Bacillus smegmatis

[0046] 1. Treat the sample to be tested with 4% NaOH solution (sample solution: NaOH solution = 1:4) at room temperature for 20 minutes, and centrifuge to remove the supernatant;

[0047] 2. Add the enrichment solution (7g of Michaelis 7H9 medium, CaCl 2 2g, 50g of ampicillin, 50g of amphotericin B, 200g of BSA, 15g of glucose, 2g of oleic acid and 0.2g of catalase were dissolved in 1L of water) 2ml, cultured at 37°C for 18-24 hours;

[0048] 3. Take 0.9ml of the above cultured bacterial solution, and add the concentration of 10 9 0.1ml of D29 phage solution per ml, incubate at 37°C for 2 hours to allow the phages to infect Mycobacterium tuberculosis;

[0049] 4. Add 0.1ml of killing agent (8% ferrous ammonium sulfate) to the above solution, and act at room temperature for 10 minutes to inactivate all bacteriophages that have not invaded Mycobacte...

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PUM

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Abstract

The invention discloses an antibody combined with a tubercle bacillus bacteriophage tail protein. The antibody can identify the tubercle bacillus bacteriophage. Besides, the invention also discloses a preparation method of the antibody, comprising the following steps of: synthesizing a tubercle bacillus bacteriophage tail protein segment, immunizing a mice by the purified tubercle bacillus bacteriophage tail protein segment, fusing the pulpar cell and with the myeloma cell, selecting cell strain with good antibody, injecting cell strain into the mice enterocoelia, and extracting ascetic fluid and purifying the antibody ten days after injection. Besides, the invention also discloses an application of the antibody in detecting tubercle bacillus. The antibody can rapidly identify the bacteriophage, further shorten the time for detecting the tubercle bacillus by a bacteriophage amplification method and improve the detection sensitivity.

Description

technical field [0001] The present invention relates to an antibody, in particular to an antibody binding to the tail protein of Mycobacterium tuberculosis phage; in addition, the present invention also relates to the preparation method and application of the antibody binding to the tail protein of Mycobacterium tuberculosis phage. Background technique [0002] Tuberculosis is a zoonotic chronic infectious disease caused by Mycobacterium tuberculosis, and is currently the main factor of death caused by a single infectious bacterium. In recent years, with the prevalence of AIDS, the increase of patients with various malignant tumors and the emergence of multi-drug resistant strains, the difficulty in the prevention and treatment of tuberculosis infection has increased. Statistics show that there are 2 billion tuberculosis infections in the world (accounting for 1 / 3 of the total population), and one person is infected every second. The global tuberculosis bacteria infects 8 mi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/08C12N5/20C12N15/06C12P21/08G01N33/569G01N33/558
Inventor 胡志能杨挥
Owner ABBOTT DIAGNOSTICS (SHANGHAI) CO LTD
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