ELISA (Enzyme Linked Immuno-Sorbent Assay) knit based on Nsp10 protein in PRRSV (porcine reproductive and respiratory syndrome virus)

A kit and protein technology, applied in the field of kits, can solve problems such as economic losses

Inactive Publication Date: 2011-07-20
GUIZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, if the pigs are immunized without distinguishing the PRRS inactivated vaccine immunized p

Method used

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  • ELISA (Enzyme Linked Immuno-Sorbent Assay) knit based on Nsp10 protein in PRRSV (porcine reproductive and respiratory syndrome virus)
  • ELISA (Enzyme Linked Immuno-Sorbent Assay) knit based on Nsp10 protein in PRRSV (porcine reproductive and respiratory syndrome virus)
  • ELISA (Enzyme Linked Immuno-Sorbent Assay) knit based on Nsp10 protein in PRRSV (porcine reproductive and respiratory syndrome virus)

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Embodiment 1 of the present invention: ELISA kit based on Nsp10 protein in PRRSV, it comprises 100 μ L concentration is the recombinant protein coating of 7.37 μ g / mL, 100 μ L positive control (positive PRRSV isolated virus is injected in unimmunized pig and prepares positive serum), 100 μL negative control (the negative control uses serum that has not been immunized and not infected with PRRSV virus) (the dilution ratio of positive control and negative control is 1:40), 100 μL blocking solution (blocking solution is skim milk, the concentration is 5g / 100 mL), 100 μL tetramethylbenzidine as the substrate solution, 100 μL sulfuric acid with a concentration of 2 mol / L as the stop solution, and 100 μL enzyme-labeled secondary antibody at a dilution of 1:1000.

Embodiment 2

[0072] Embodiment 2 of the present invention: based on the ELISA kit of Nsp10 protein in PRRSV, it includes 50 μ L of recombinant protein coating with a concentration of 59 μ g / mL, 50 μ L of positive control (positive PRRSV isolated virus is injected into unimmunized pigs and prepared) Positive serum), 50 μL negative control (the negative control uses serum that has not been immunized or infected with PRRSV virus) (the dilution ratio of positive control and negative control is 1:10), 50 μL blocking solution (blocking solution is BSA, the concentration is 1g / 100mL ), 50 μL heterocyclic azine as substrate solution, 50 μL hydrochloric acid with a concentration of 1 mol / L as stop solution, and 50 μL enzyme-labeled secondary antibody at a dilution of 1:500.

Embodiment 3

[0073] Embodiment 3 of the present invention: ELISA kit based on Nsp10 protein in PRRSV, it includes 200 μ L concentration is the recombinant protein coating of 0.922 μ g / mL, 200 μ L positive control (positive PRRSV isolated virus is injected in unimmunized pig and prepares positive serum), 200 μL negative control (the negative control uses serum that has not been exempted or not infected with PRRSV virus) (the dilution ratio of positive control and negative control is 1:320), 200 μL blocking solution (blocking solution is gelatin, the concentration is 1g / 100mL ), 200 μL o-phenylenediamine as the substrate solution, 50 μL sulfuric acid with a concentration of 2 mol / L as the stop solution, and 200 μL enzyme-labeled secondary antibody at a dilution of 1:2000.

[0074] When mixing the above-mentioned BSA, skim milk, gelatin or FBS, first weigh the corresponding materials according to the predetermined weight, place them in 90mL sterilized deionized water for dissolution, and then ...

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Abstract

The invention discloses an ELISA (Enzyme Linked Immuno-Sorbent Assay) knit based on Nsp10 protein in PRRSV (porcine reproductive and respiratory syndrome virus), which comprises 50-200 ml of recombined protein peridium, 50-200 ml of positive reference, 50-200 ml of negative reference, 50-200 ml of sealing liquid, 50-200 ml of enzyme labeled secondary antibody, 50-200 ml of substrate liquid and 50-200 ml of stop solution. The purified Nsp10 recombined protein is taken as envelope antigen, and an ELISA method is established to optimize the reaction condition and research an ELISA knit based on Nsp10 protein in PRRSV; via comparing the ELISA detection result with the positive reference and the negative reference, a detection conclusion is obtained; a detection conclusion is obtained through the comparison between the ELISA detection result and the negative reference and the positive reference; the conclusion can basically distinguish PRRS inactivated vaccine immune swinery from inapparent infected swinery; therefore, reference data is provided for the clinical diagnosis and the prevention and control of the PRSS; according to the detection result, the uninfected swinery is selectively immune; therefore, the immune aimlessness can be reduced, and the economic loss is effectively avoided. The ELISA knit has the characteristics of good specificity, sensitivity, repeatability, and the like, and is good in use effect.

Description

technical field [0001] The invention relates to a kit, especially an ELISA kit based on Nsp10 protein in PRRSV. Background technique [0002] Porcine reproductive and respiratory syndrome (PRRS) is a serious infectious disease characterized by reproductive disorders such as abortion, premature birth, and weak stillbirths in sows, dyspnea in piglets, high mortality, and respiratory symptoms in fattening pigs. It is one of the important pathogens of the pig industry, which has caused huge economic losses to the pig industry in my country. Because PRRSV has the phenomenon of antibody-dependent infection enhancement, it can be seen clinically that PRRS symptoms in pigs will be aggravated after PRRS vaccination, and the antibodies induced by the vaccine will enhance the replication of wild virus in pigs. Non-structural proteins are produced during virus replication, and can only be produced when the body is infected by the virus. Inactivated vaccine immunization only produces ant...

Claims

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Application Information

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IPC IPC(8): G01N33/68
Inventor 周碧君周思旋文明王开功程振涛岳筠刘忠伟汪德生温贵兰
Owner GUIZHOU UNIV
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