Method of detecting specific killing effects of chicken spleen CD8<+>T cells based on flow cytometry

Abstract

The invention relates to a method of detecting specific killing effects of chicken spleen CD8<+>T cells based on flow cytometry. The method comprises a first step of bacterial culture in which the bacteria are salmonella pullorum S06004 and S06004deltaspiC, a second step of chick immunization and sampling, a third step of target cell-macrophage preparation, a fourth step of effector cell-chicken spleen CD8<+>T cell preparation which comprises a substep of preparation of a chicken spleen lymphocyte suspension and a substep of magnetic bead sorting of a chicken spleen CD8<+>T cell and a fifth step of specifically killing macrophage by CTL. Compared with the prior art, the method disclosed in the invention has the following beneficial effects that the macrophage (target cell) obtained by adherence has high purity, the sorting effects for chicken spleen CD8<+>T cells are obvious, and the separation time is short; with the salmonella host cell- macrophages as the target cell, the salmonellaremoving ability by the host can be reflected more accurately; and the flow cytometry is used for chicken spleen CD8<+>T cell specific killing effects and can detect the CTL effects of the chicken spleen CD8<+>T cell in a single cell level.

Description

technical field [0001] The invention relates to a method for detecting chicken spleen CD8 based on flow cytometry + Methods of T cell-specific killing. Background technique [0002] Cellular immune response is mediated by T lymphocytes, which is conducive to promoting the production of antibodies in the body, and also helps to remove pathogenic microorganisms (such as bacteria, viruses, etc.) parasitic in the cells. In addition, cellular immune response also plays an important role in poultry immune surveillance and anti-tumor (such as Marek's disease) immunity. After poultry is infected by pathogenic microorganisms, T cells play a vital role in the immune response process by producing pro-inflammatory cytokines, regulating local and systemic immune responses, activating macrophages, and clearing intracellular bacteria. Among them, cytotoxic T cells (CTL) have lytic activity and play a key role in the recognition and removal of abnormal cells (such as tumors and pathogen-i...

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Problems solved by technology

[0005] The object of the present invention is to target radioactive chromium ( 51 Cr) release assay cytotoxicity method has the disadvantages of low sensitivity and strong radioactivity, and establishes a newer, more reliable method that can detect cytotoxicity at the single-cell level (i.e., a method based on flow cytometry to detect chicken spleen CD8 + T cell-specific killing method)

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