ELISA (enzyme-linked immuno sorbent assay) detection kit of porcine pseudorabies virus IgM antibody

A porcine pseudorabies virus, detection kit technology, applied in the direction of viral peptides, measuring devices, recombinant DNA technology, etc., can solve problems such as antibodies that cannot detect specific antigens

Inactive Publication Date: 2013-09-18
WUHAN CHOPPER BIOLOGY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are many kits for PRV antibody detection in the foreign market, but these kits cannot detect antibodies against specific antigens, that is, detect antibodies against what kind of pathogens, such as porcine pseudorabies virus gE, gB and gD protein antigens or other viruses, etc.

Method used

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  • ELISA (enzyme-linked immuno sorbent assay) detection kit of porcine pseudorabies virus IgM antibody
  • ELISA (enzyme-linked immuno sorbent assay) detection kit of porcine pseudorabies virus IgM antibody
  • ELISA (enzyme-linked immuno sorbent assay) detection kit of porcine pseudorabies virus IgM antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] Example 1 Preparation method of porcine pseudorabies virus (PRV) IgM antibody ELISA detection kit

[0063] 1. The porcine pseudorabies virus (PRV) IgM monoclonal antibody of the present embodiment was purchased from MP Biomedicals, Incorporated;

[0064] 2. the preparation method of horseradish peroxidase (HRP)-anti-porcine pseudorabies virus gE, gB and gD protein antibody enzyme conjugate comprises the steps:

[0065] 1) The purified gE, gB and gD proteins were routinely immunized rabbits respectively, and when the ELISA titer of the rabbit serum reached above 1:1000, the serum was taken;

[0066] 2) Precipitating and purifying the serum prepared in step 1) through ammonium sulfate to obtain purified serum;

[0067] 3) Carry out HRP labeling to the purified serum prepared in step 2) by an improved sodium periodate oxidation method to prepare horseradish peroxidase (HRP)-anti-gE, gB and gD protein antibody enzyme conjugates;

[0068] 4) Add 0.1-10% bovine serum albu...

Embodiment 2

[0083] Example 2 Detection method of porcine pseudorabies virus IgM antibody ELISA detection kit

[0084] Utilize the detection method of porcine pseudorabies virus IgM antibody ELISA detection kit of the present invention, comprise the steps:

[0085] 1) Dilute the sample to be tested 100 times with the sample diluent, add it to the wells of the microplate plate evenly coated with porcine IgM monoclonal antibody, add 100 μl to each well, and set a positive control group, a negative control group and a blank control group at the same time 100 μl positive control solution was added to the positive control group, 100 μl negative control solution was added to the negative control group, 100 μl sample diluent was added to the blank control group, and 3 wells were added in parallel for each sample and control;

[0086] 2) After adding the samples, incubate the ELISA plate at 37° C. for 1 hour, wash the plate with washing solution 3 times, and spin dry. The washing solution is pre...

Embodiment 3

[0092] Example 3 Specificity and sensitivity test of the kit of the present invention

[0093] 1. Specific detection

[0094] According to the detection method described in Example 2, the kits prepared in Example 1 were used to detect respectively the early serum of porcine pseudorabies live vaccine immunization, the early serum of porcine PRV infection, the purified porcine PRV-gE IgG antibody, and the purified PRV-gB IgG antibody, purified PRV-gD IgG antibody, porcine PCV2 infection early serum, porcine PRRS virus antibody positive serum, swine fever virus antibody positive serum and porcine PRV negative serum.

[0095] Dilute the sample to be tested 100 times with the sample diluent, add it to the wells of the microplate plate uniformly coated with porcine IgM monoclonal antibody, add 100 μl to each well, set up a positive control group, a negative control group and a blank control group in parallel, Among them, 100 μl positive control solution was added to the positive ...

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Abstract

The invention relates to an ELISA (enzyme-linked immuno sorbent assay) detection kit of a porcine pseudorabies virus IgM antibody, and a preparation method and application thereof. The ELISA detection kit of the porcine pseudorabies virus IgM antibody disclosed by the invention comprises an elisa plate of enveloping an IgM monoclonal antibody, sealing liquid, sample diluting liquid, detecting antigen, enzyme conjugate, concentrated scrubbing solution, enzyme substrate A solution, enzyme substrate B solution and stop buffer; the detecting antigen is prepared from purified porcine pseudorabies virus gE protein, structure protein gB and structure protein gD; and the enzyme conjugate is horse radish peroxidase-anti-gE, gB or gD protein enzyme compound. The specificity of the kit disclosed by the invention can be up to 100%; the sensitivity is 1:640; and the kit can be used for early diagnosis of porcine pseudorabies virus infection.

Description

technical field [0001] The invention relates to a detection kit, in particular to a porcine pseudorabies virus IgM antibody ELISA detection kit, a detection method and an application thereof. Background technique [0002] Pseudorabies is an acute infectious disease caused by pseudorabies virus (PRV), including a variety of livestock and wild animals. Pigs are the main storage host and source of infection for pseudorabies. Healthy pigs can be infected with the disease by direct contact with sick pigs and infected pigs. Adult pigs are often insidiously infected; infected pregnant sows have symptoms such as miscarriage, stillbirth, weak fetus, and mummified fetuses; infected newborn piglets have fever and neurological symptoms, and even collapse and die, with a mortality rate of up to 100%. [0003] Since pseudorabies was first discovered in the United States in 1902, the disease has spread worldwide. Since the first discovery of pseudorabies in my country in 1947, 31 provin...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577C12N15/38C12N15/70C07K14/03
Inventor 廖园园漆世华孙庆歌李建秦伟谢红玲温文生
Owner WUHAN CHOPPER BIOLOGY
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