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Kit for testing neutralizing antibody racing ELISA in human and animal rabies

A detection kit and kit technology, applied in the field of biological vaccine preparation, can solve problems such as long time required, poor sensitivity and specificity, large difference in antibody potency, etc., and achieve accurate quantitative determination, good consistency, and time-consuming short effect

Inactive Publication Date: 2008-08-27
MILITARY VETERINARY RES INST PLA MILITARY MEDICAL ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the two can accurately measure the level of rabies neutralizing antibody, both of them use live cells and viruses to measure, the operation is complicated, it takes a long time, and there are many influencing factors
In recent years, a variety of rabies neutralizing antibody detection technologies have been developed successively, such as flow cytometry, indirect ELISA technology, and chromatography diagnostic test strip technology, etc. Compared with the above two recommended methods, these technologies have higher sensitivity and specificity The results are poor, and the obtained antibody potency values ​​vary greatly, and cannot be accurately quantified, especially for the antibody level determination effect of the neutralizing antibody critical value (0.5IU / ml), it is difficult to make an accurate determination

Method used

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  • Kit for testing neutralizing antibody racing ELISA in human and animal rabies
  • Kit for testing neutralizing antibody racing ELISA in human and animal rabies

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Preparation method of the kit (1)

[0038] 1. Preparation and Coating of Rabies Virus

[0039] Rabies virus was inoculated into BHK-21 monolayer cells, cultured for 3 to 4 days, frozen and thawed twice, cells were lysed, and culture medium was harvested. When coating, dilute the virus with the coating solution to a total protein concentration of 100 μg / ml, add 100 μl to the enzyme-linked plate, and coat overnight at 4°C; rinse three times with PBS-Tween 20 buffer, and wash the virus three times in each well. Add 100 μl of blocking solution (containing 5% skimmed milk powder and 0.01% thimerosal), place at 37°C for 1 hour, discard the blocking solution, seal it in a vacuum-sealed aluminum foil bag, and store at 4°C.

[0040] 2. Preparation of standard serum

[0041] 2.1 Preparation of human standard serum

[0042]Commercially available human rabies vaccine (Dalian Jingang Andy Biological Products Co., Ltd.) was purchased, and the volunteers were immunized three times ...

Embodiment 2

[0067] The preparation method of the kit (2)

[0068] 1. Preparation and Coating of Rabies Virus

[0069] Rabies virus was inoculated into BHK-21 monolayer cells, cultured for 3 to 4 days, frozen and thawed twice, cells were lysed, and culture medium was harvested. When coating, dilute the virus with the coating solution to a total protein concentration of 100 μg / ml, add 100 μl to the enzyme-linked plate, and coat overnight at 4°C; rinse three times with PBS-Tween 20 buffer, and wash the virus three times in each well. Add 100 μl of blocking solution (containing 5% skimmed milk powder and 0.01% thimerosal), place at 37°C for 1 hour, discard the blocking solution, seal it in a vacuum-sealed aluminum foil bag, and store at 4°C.

[0070] 2. Preparation of standard serum

[0071] 2.1 Preparation of human standard serum

[0072] Commercially available human rabies vaccine (Dalian Jingang Andy Biological Products Co., Ltd.) was purchased, and the volunteers were immunized three t...

Embodiment 3

[0098] The preparation method of the kit (3)

[0099] 1. Preparation and Coating of Rabies Virus

[0100] Rabies virus was inoculated into BHK-21 monolayer cells, cultured for 3 to 4 days, frozen and thawed twice, cells were lysed, and culture medium was harvested. When coating, dilute the virus with the coating solution to a total protein concentration of 100 μg / ml, add 100 μl to the enzyme-linked plate, and coat overnight at 4°C; rinse three times with PBS-Tween 20 buffer, and wash the virus three times in each well. Add 100 μl of blocking solution (containing 5% skimmed milk powder and 0.01% thimerosal), place at 37°C for 1 hour, discard the blocking solution, seal it in a vacuum-sealed aluminum foil bag, and store at 4°C.

[0101] 2. Preparation of standard serum

[0102] 2.1 Preparation of human standard serum

[0103] Commercially available human rabies vaccine (Dalian Jingang Andy Biological Products Co., Ltd.) was purchased, and the volunteers were immunized three t...

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Abstract

The invention discloses a reagent box for detecting hydrophobia neutralizing antibody competition ELISA of human beings and animals, wherein the reagent box can easily, quickly, accurately and quantitatively detect the hydrophobia neutralizing antibody in blood serums of human beings and animals by marking the hydrophobia neutralizing antibody, the standard serum and the envelope antigen. By using hydrophobia virosome or virus glycoprotein to coat enzyme synapticulae, the enzyme labeling hydrophobia neutralizing antibody is mixed with the blood serum to be tested and the standard serum respectively according to a certain ratio and reacts with the hydrophobia virus glycoprotein antigen coated on the enzyme synapticulae, a standard curve is drawn according to the OD value of the standard blood serum reaction and the known neutralizing titer after the color development, and the titer of the corresponding neutralizing antibody is obtained from the standard curve according to the OD value of the reaction of the blood serum to be tested. The reagent box has the advantages of accurately and quantitatively detecting the neutralizing antibody of the hydrophobia virus, along with simple operation and short time; moreover, the test result of the invention keeps a good consistence with test results of neutralizing test methods recommended by WHO and OIE.

Description

Technical field: [0001] The invention discloses a human and animal rabies neutralizing antibody competition ELISA detection kit, which accurately and quantitatively detects the titer of rabies neutralizing antibody in human and animal serum through the competition reaction of enzyme-labeled antibody and serum antibody, and belongs to the technical field of biological vaccine preparation . Background technique: [0002] Rabies is an important zoonotic infectious disease. After rabies infects humans or animals, the case fatality rate is 100%. Active rabies prophylaxis in animals and post-exposure treatment (vaccine and immunoglobulin injections) in rabies-exposed populations are key to preventing rabies. Rabies vaccine can induce five kinds of antibodies against its structural proteins in the body, among which the neutralizing antibody against glycoprotein is currently the only recognized anti-infection antibody. The World Health Organization (WHO) and the Animal World Healt...

Claims

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Application Information

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IPC IPC(8): G01N33/53
Inventor 扈荣良刘晔张守峰张菲
Owner MILITARY VETERINARY RES INST PLA MILITARY MEDICAL ACAD OF SCI
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