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Cytomegalovirus specificity cellular immunity response external detecting model

A cytomegalovirus and cellular immunity technology, applied in the field of in vitro detection models of cytomegalovirus-specific cellular immune responses, can solve the problem of reduced expression of MHC class I molecules on the surface of infected cells, failure to detect virus-specific cellular immune responses, and inability to detect virus-specific cellular immune responses. Appropriate detection models, etc.

Inactive Publication Date: 2008-08-20
王延军 +2
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Problems solved by technology

Infect human fibroblasts with classic CMV laboratory strains as antigen-presenting cells, and detect the expression of cytokines and HLA molecules by flow cytometry, and analyze the ability of the cells infected by the virus strain to induce immune responses. The study found that the infected cells The expression of MHC class I molecules on the surface is significantly reduced, and the virus-specific cellular immune response cannot be detected, so it is not suitable as a detection model

Method used

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Embodiment Construction

[0004] 1. CMV infection of human fibroblasts to stimulate peripheral blood lymphocytes: Seed human fibroblasts in a 24-well cell culture plate, add 2ml DMEM medium to each well, the cells grow into a single layer, cover the whole hole, follow the 1:1 ratio (quantity ratio), add CMV mutant virus liquid, infect at 37°C for 1 hour, discard free virus liquid, add 2ml DMEM cell culture medium, incubate at 37°C, 5% CO 2 Incubated in an incubator for 48 hours, washed once with phosphate buffered saline (PBS), and then added 2×10 6 Peripheral blood lymphocytes were incubated for 2 hours, then added with 10 μg / ml Golgi inhibitor Brefeldin A, and cultured for 4 hours.

[0005] 2. Detection of IFN-γ by intracellular cytokine labeling method: after the culture, collect the cells, wash once with PBS, resuspend the cells in PBS solution containing 0.02% EDTA, incubate at 37°C for 15 minutes, centrifuge, wash once with PBS, Resuspend cells in erythrocyte lysate and incubate at room temperat...

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Abstract

The invention provides a test for new anti-cytomegalovirus medicine efficacy, establishing a testing model in vitro of cytomegalovirus specific immune response, relative reports have not been seen at home and abroad nowadays. It is because that the virus suppresses the expression of major histocompatibility complex-I and reflects antigen-presenting in model established with cells infected by traditional cytomegalovirus laboratory strain, virus-specific immune response can not be tested. The invention uses cytomegalovirus to infect human fibroblasts cells as the antigen for presenting cells, and stimulates target cells, flow cytometry is used to test expression of cytokines gamma-interferon which is used as indicator for establishing test model in vitro of cytomegalovirus specific immune response. Adding test medicine into the model can test the effect of the medicine to virus-specific immune response.

Description

technical field [0001] The invention relates to a model capable of detecting cytomegalovirus (CMV)-specific cellular immune response in vitro. The model can not only conveniently detect the cytomegalovirus-specific cellular immune response of patients in vitro, but also screen new drugs. Background technique [0002] In recent years, scientists have devoted themselves to developing new anti-CMV drugs in order to inhibit the immune pathological damage caused by the virus. To detect the efficacy of new drugs, it is first necessary to establish a CMV-specific immune response in vitro detection model, but there are no related reports at home and abroad. Infect human fibroblasts with classic CMV laboratory strains as antigen-presenting cells, and detect the expression of cytokines and HLA molecules by flow cytometry, and analyze the ability of the cells infected by the virus strain to induce immune responses. The study found that the infected cells The expression of MHC class I ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569
Inventor 王延军孙斌武志明
Owner 王延军
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