Application of group of serum differential proteins in preparation of reagent for detecting toxoplasma gondii infection of pigs and detection kit for identification

A technology of differential protein and Toxoplasma gondii, applied in biological testing, measuring devices, material inspection products, etc., can solve the problems of low detection rate, low specificity and sensitivity, time-consuming and labor-intensive, and achieve accurate identification and large market. Foreground effect

Inactive Publication Date: 2020-12-18
JIANGSU AGRI ANIMAL HUSBANDRY VOCATIONAL COLLEGE
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  • Summary
  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

The disease has a high morbidity and fatality rate in domestic animals such as pigs. The routine diagnosis is mainly based on laboratory tests, including pathogens and serological tests. The pathogenic test is mainly based on animal visceral tissue smears or abdominal cavity inspection after animal inoculation. Liquid worms, confirmed after pathogens are found, this method is stable and reliable, but time-consuming and laborious, the detection rate is low; serological methods mainly include enzyme-linked immunosorbent assay (ELISA), indirect hemagglutination assay (IHA) and indirect immunofluorescence antibody test (IFA), PCR detection, etc., are simple and fast to operate, but the specificity and sensitivity are not high, and false positives are prone to occur

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0022] 1. Detection of differentially expressed proteins between pigs infected with Toxoplasma gondii and healthy controls by iTRAQ technology

[0023] 1. Test samples

[0024] 20 cases of Toxoplasma gondii-infected pig serum and 20 healthy pig serum from a certain place in Jiangsu. Collect 2 mL of whole blood on an empty stomach in the morning, let it stand at 4°C for 1-2 hours until the blood coagulates and precipitate the serum, centrifuge at 3000g for 10 minutes, collect the supernatant, aliquot it on ice and store it at -80°C for later use.

[0025] 2. Detection method

[0026] (1) Use the Proteominer kit to remove high-abundance proteins in serum: add DTT (dithiothreitol) at a final concentration of 10 mM to the enriched protein sample, and bathe in water at 56°C for 1 hour; after cooling to room temperature, add the final concentration 55mM IAM (iodoacetamide), placed in a dark room for 45min; add 1mL of cold acetone to precipitate overnight, centrifuge at 25000rpm at...

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Abstract

The invention discloses application of a group of serum differential proteins in preparation of a reagent for detecting toxoplasma gondii infection of pigs. The serum differential proteins are one ora combination of more than one of fragments of toxoplasma gondii surface membrane proteins, HF proteins, vacuolar proteins, gelsolin and apolipoprotein CI. Whether the serum differential protein exists in serum or not is detected through an iTRAQ technology, and the concentration variation of the differential protein is detected and verified in combination with an MRM technology. The invention also discloses a detection kit for identifying toxoplasma gondii infection of pigs. The detection kit comprises an antibody for detecting the serum differential protein, a phosphate buffer solution and an ELISA 96-well plate. Whether pigs are infected with toxoplasma gondii or not can be rapidly and accurately identified, and the serum differential protein provided by the invention has the characteristics of objectivity, specificity and accuracy when used as a toxoplasmosis detection marker, and has a great market prospect.

Description

technical field [0001] The invention belongs to the field of diagnostic protein markers for toxoplasma gondii infection in pigs, and specifically relates to the application of a group of serum differential proteins in the preparation of reagents for detecting toxoplasma gondii infection in pigs and a detection kit for identification. Background technique [0002] Toxoplasma gondii (Toxoplasma gondii) is an obligate intracellular parasitic opportunistic pathogenic protozoan, which can spread in various animals such as cattle, sheep, pigs, chickens, cats, etc. The focus is on strengthening disease diagnosis, and there are no ideal drugs and vaccines yet. The disease has a high morbidity and fatality rate in domestic animals such as pigs. The routine diagnosis is mainly based on laboratory tests, including pathogens and serological tests. The pathogenic test is mainly based on animal visceral tissue smears or abdominal cavity inspection after animal inoculation. Liquid worms, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/569G01N27/62
CPCG01N33/6848G01N33/56905G01N2333/45
Inventor 郝福星黄玉政袁橙窦浩华游淑惠
Owner JIANGSU AGRI ANIMAL HUSBANDRY VOCATIONAL COLLEGE
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