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Composition for detecting mycoplasma antibody and application thereof

A technology for mycoplasma and composition, applied in the field of compositions for detecting mycoplasma antibodies, can solve problems such as false positive test results, and achieve the effects of easy acquisition and low false positive rate

Active Publication Date: 2016-05-04
JIANGSU ACAD OF AGRI SCI
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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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  • Composition for detecting mycoplasma antibody and application thereof
  • Composition for detecting mycoplasma antibody and application thereof
  • Composition for detecting mycoplasma antibody and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1 Preparation of Mycoplasma hyorrhinos antigenic peptide

[0021] 1. Selection of antigenic peptides of M. hyorhinosa

[0022] In the present invention, a large amount of comparison work is performed on the whole sequence of Mycoplasma hyorrhinosus, and a suitable protein with high specificity is screened as an antigen for detecting Mycoplasma hyorrhinosus antibody. Finally, it was found that some polypeptides of the variable lipoprotein (vlp) family on the surface of Mycoplasma hyorrhinos could be used as antigenic peptides for the specific detection of Mycoplasma hyosinus antibodies, and would not be interfered by Mycoplasma hyopneumoniae, Mycoplasma hyos synovium or Mycoplasma flocculents. The vlp family consists of 7 members, namely vlpA, vlpB, vlpC, vlpD, vlpE, vlpF, vlpG. The gene coding regions of vlp members have the same structure and are divided into three parts: Ⅰ region, Ⅱ region and Ⅲ region. Among them, region III encodes a repeating peptide segm...

Embodiment 2

[0037] Example 2 Preparation of Mycoplasma hyorrhinos antigenic peptide composition

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

Embodiment 3

[0039] Example 3 The specific method for detecting the serum antibody of Mycoplasma hyorrhinosus by indirect ELISA method with the antigenic peptide composition of Mycoplasma hyorrhinosus as the coating antigen

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

[0041] PBS buffer (concentration is 0.01M, pH7.2): take 8gNaCl, 0.2gKCl, 1.44gNa 2 HPO 4 and 0.24gKH 2 PO 4 , dissolve in 800ml of distilled water, adjust the pH to 7.2 with hydrochloric acid, and finally add distilled water to dilute to 1L.

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

[0043] Na 2 CO 3 -NaHCO 3 Buffer solution (concentration is 0.05M, pH9.6): take 1.59gNa 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 Biologi...

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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 its application. Background technique [0002] Mycoplasma hyorhinis, a member of the genus Mycoplasma of 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 hyorrhinos is a common pathogen in clinical pig farms, usually transmitted from sows or large pigs to young pigs, usually by upper respiratory tract infection through droplets or direct contact. Once infected pigs, the mycoplasma spreads rapidly in the upper respiratory tract and can be isolated from the lungs and nasopharyngeal tube of infected pigs, and then can migrate to the whole body through the respiratory tract. Mycoplasma hyorhinosa can cause porcine polyserositis, arthritis, otitis media, pneumonia and other diseases. ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K19/00
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 ACAD OF AGRI SCI
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