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Application of porcine circovirus type 4-specific antigen in the preparation of a kit for detecting porcine circovirus type 4 antibody and the kit

A porcine circovirus and kit technology, which is applied in the field of immune detection, can solve problems such as lack of accurate virus detection methods, and achieve the effects of good clinical application value, high sensitivity and high throughput

Active Publication Date: 2022-04-19
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is currently a lack of an on-site rapid and accurate detection method for the virus

Method used

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  • Application of porcine circovirus type 4-specific antigen in the preparation of a kit for detecting porcine circovirus type 4 antibody and the kit
  • Application of porcine circovirus type 4-specific antigen in the preparation of a kit for detecting porcine circovirus type 4 antibody and the kit
  • Application of porcine circovirus type 4-specific antigen in the preparation of a kit for detecting porcine circovirus type 4 antibody and the kit

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preparation example Construction

[0015] The present invention also provides a test kit for detecting porcine circovirus type 4 antibody, said kit comprising: an ELISA plate coated with porcine circovirus type 4 specific antigen, a negative control, a positive control, and sample dilution solution, blocking solution, washing solution, enzyme-labeled secondary antibody solution, chromogenic solution and reaction termination solution; the amino acid sequence of the porcine circovirus type 4 specific antigen is shown in SEQID NO.1. In the present invention, the kit preferably also includes an ELISA coating solution, which can effectively activate the antigen and fully expose the antigenic determinant recognized by the antibody. The preparation method of the ELISA coating solution includes: NaCO 3 1.59g, NaHCO 3 2.93 g was dissolved in 1 L of distilled water, and the pH was adjusted to 9.6. In the present invention, the preparation method of the washing liquid preferably includes: taking 8g of NaCl, KH 2 PO 4 0...

Embodiment 1

[0020] Screening and preparation of PCV4 specific antigen

[0021] 1PCV4 type-specific antigen screening.

[0022] By comparing the amino acid sequences of different circovirus genotypes, the amino acid sequence of GHYDFKLKDFIPKGPGTIVNLYSLPFA was screened. This peptide has the lowest amino acid sequence homology with PCV1, PCV2, and PCV3, and has good antigenic reactogenicity.

[0023] Preliminary establishment of 2PCV4 indirect ELISA method

[0024] 2.1 Determination of antigen coating concentration

[0025] Dilute PCV4 antigen with coating solution to protein content of 5 μg / mL, 10 μg / mL, 20 μg / mL, 40 μg / mL respectively, add to 96-well ELISA plate, add 100 μL to each well, and place overnight at 4 °C. The next day, shake off the liquid in the well, wash with 300 μL washing solution twice, 3 minutes each time, and pat dry.

[0026] 2.2 Closed

[0027] Add blocking solution to the microtiter plate, 100 μL per well, and react at 37°C for 30 minutes.

[0028] 2.3 Add primar...

Embodiment 2

[0060] Comparison of PCV4 indirect ELISA method and PCV4 real-time fluorescent PCR method

[0061] The purpose is to verify the coincidence rate of PCV4 indirect ELISA method and PCV4 real-time fluorescent PCR method for detection of clinical samples. Take 10 clinical serum samples and 10 negative samples tested positive by PCV4 real-time fluorescent PCR method, and use PCV4 indirect ELISA kit for detection.

[0062] Table 2 Comparison results of PCV4 indirect ELISA method and PCV4 real-time fluorescent PCR method

[0063]

[0064]

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Abstract

The invention relates to an application of a porcine circovirus type 4-specific antigen in the preparation of a kit for detecting porcine circovirus type 4 antibody and the kit, and belongs to the technical field of immune detection. The present invention provides the application of a porcine circovirus type 4 specific antigen in the preparation of a kit for detecting porcine circovirus type 4 antibody, the amino acid sequence of the porcine circovirus type 4 specific antigen is as shown in SEQ ID NO.1 shown. The kit prepared by the application of the invention has good detection specificity, high throughput, has important clinical application value, and has rapid and accurate detection, which can realize the epidemiological detection of PCV4 in pig farms, and can also be used for vaccine immunization Evaluation of PCV4 antibodies in porcine serum.

Description

technical field [0001] The invention relates to the technical field of immune detection, in particular to the application of a porcine circovirus type 4 specific antigen in the preparation of a kit for detecting porcine circovirus type 4 antibody and the kit. Background technique [0002] Porcine circovirus (PCV) is a circular, single-stranded DNA virus with a genome. According to the characteristics of the genome, four genotypes of PCV1, PCV2, PCV3 and PCV4 have been found. Among them, PCV1 can be detected in pig herds, but it does not cause disease; PCV2 is the main pathogen that causes porcine post-weaning failure syndrome; PCV3 is a virus associated with sow dermatitis syndrome and reproductive disorders; the newly discovered pig Circovirus type 4 (PCV4) may cause various diseases of porcine respiratory tract and digestive tract, and may be closely related to porcine dermatitis and nephrotic syndrome. At present, the virus has been reported in many provinces in my count...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/569
CPCG01N33/56983G01N2469/20G01N2333/01
Inventor 李向东刘金彪陈昌海范娟
Owner YANGZHOU UNIV