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Combinant for detecting mycoplasma antibody and application of combinant

A technology of mycoplasma and composition, which is applied in the field of composition for detecting mycoplasma antibodies, can solve problems such as false positive detection results, and achieve the effect of easy acquisition and low false positive rate

Active Publication Date: 2015-06-24
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Recently, it has been reported in the literature that the ELISA method was initially established to detect serum antibodies using the Mycoplasma hyorhina P37 protein as the coating antigen. However, in fact, through sequence comparison, it can be found that the P37 protein has obvious cross-epitopes with a variety of other Mycoplasma hyopneumoniae, especially Mycoplasma hyopneumoniae. , can lead to serious false positive detection results

Method used

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  • Combinant for detecting mycoplasma antibody and application of combinant
  • Combinant for detecting mycoplasma antibody and application of combinant
  • Combinant for detecting mycoplasma antibody and application of combinant

Examples

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Embodiment 1

[0020] The preparation of embodiment 1 mycoplasma hyorhina antigenic peptide

[0021] 1. Selection of Mycoplasma hyorhini antigenic peptides

[0022] In the invention, a large amount of comparison work is carried out on the whole sequence of mycoplasma hyorhina, and a suitable protein with high specificity is screened as an antigen for detecting the mycoplasma hyorhina antibody. Finally, it was found that some polypeptides of the variable lipoprotein (vlp) family on the surface of M. hyorhina can be used as antigenic peptides for the specific detection of M. hyorhina antibodies without being interfered by M. hyopneumoniae, M. hyosynovium or M. flocculus. The vlp family contains seven members, namely vlpA, vlpB, vlpC, vlpD, vlpE, vlpF, and vlpG. The gene coding regions of vlp members have the same structure and are divided into three parts: region Ⅰ, region Ⅱ and region Ⅲ. Among them, region III encodes repetitive peptides, which are composed of tandem repeats of peptides of ...

Embodiment 2

[0037] The preparation of embodiment 2 mycoplasma hyorhinois antigenic peptide composition

[0038] The aqueous solutions of 7 kinds of antigen peptide conjugates KLH-PepvlpA, KLH-PepvlpB, KLH-PepvlpC, KLH-PepvlpD, KLH-PepvlpE, KLH-PepvlpF, and KLH-PepvlpG were prepared respectively, with a concentration of 1 mg / ml. The aqueous solutions of the above seven kinds of antigen peptide conjugates were mixed to obtain the Mycoplasma hyorhina antigen peptide composition. The molar concentration of each antigen peptide conjugate in the Mycoplasma hyorhini antigen peptide composition is the same.

Embodiment 3

[0039] Example 3 Using the Mycoplasma hyorhina Antigen Peptide Composition as a Coating Antigen and Using the Indirect ELISA Method to Detect Mycoplasma Hyorhinosus Serum Antibody Specific Method

[0040] 1. The preparation method of reagent in the present embodiment:

[0041] PBS buffer solution (0.01M concentration, pH7.2): take 8g NaCl, 0.2g KCl, 1.44g Na 2 HPO 4 and 0.24g KH 2 PO 4 , dissolved in 800ml of distilled water, adjusted the pH to 7.2 with hydrochloric acid, and finally added distilled water to make up to 1L.

[0042] PBST: Add 0.05% tween-20 by volume to PBS buffer solution with a concentration of 0.01M and a pH of 7.2.

[0043] Na 2 CO 3 -NaHCO 3 Buffer (0.05M concentration, pH9.6): take 1.59g Na 2 CO 3、 2.93g NaHCO 3 , add distilled water to dissolve, and then dilute to 1000mL.

[0044] Enzyme-labeled secondary antibody: horseradish peroxidase-labeled goat anti-rabbit (or goat anti-pig) IgG, Wuhan Boster Bioengineering Co., Ltd.

[0045] Chromogeni...

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Abstract

The invention provides a combinant for detecting a mycoplasma antibody and application of the combinant, and belongs to the technical field of biology. The combinant comprises seven polypeptides, wherein amino acid sequences of the polypeptides are shown in SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:7 respectively. The combinant can be used for specifically detecting the mycoplasma antibody of a pig nose, and the cross reaction on the mycoplasma antibody and other mycoplasmas or other pathogenic infection samples which are commonly seen on the pig body is avoided, and the false positive rate is extremely low. Detection samples are easily available, and the normal feeding management of a pig farm cannot be influenced.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a composition for detecting mycoplasma antibodies and an application thereof. Background technique [0002] Mycoplasma hyorhinis, a member of the genus Mycoplasma in the family Mycoplasmataceae, was first isolated from the nasal cavity of pigs with infectious atrophic rhinitis by Carler and Mckay in 1953, so it was named Mycoplasma hyorhinis. Mycoplasma hyorhinosum is a common pathogen in clinical pig farms. It is usually transmitted from sows or older pigs to piglets, and is usually transmitted by upper respiratory tract infection through droplets or direct contact. Once pigs are infected, the mycoplasma spreads rapidly in the upper respiratory tract and can be isolated from the lungs and nasopharyngeal tubes of infected pigs, and then can migrate to the whole body through the respiratory tract. Mycoplasma hyorhini can cause porcine polyserositis, arthritis, otitis medi...

Claims

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Application Information

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IPC IPC(8): G01N33/68
CPCC07K14/30G01N33/56933G01N33/68G01N2333/30
Inventor 熊祺琰邵国青王佳刘茂军冯志新韦艳娜马庆红华利忠倪博
Owner JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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