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A method for detecting the immunosuppressive function of regulatory T cells

A cellular immune and regulatory technology, applied in biochemical equipment and methods, microbial determination/inspection, fluorescence/phosphorescence, etc., can solve the problems of radioactive pollution in the surrounding environment, and achieve the effect of simple operation, long time and high efficiency

Inactive Publication Date: 2011-12-21
顾鹏
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Problems solved by technology

[0003] This method not only requires special expensive equipment such as liquid scintillation instrument, cell harvester and special reagents such as β liquid scintillation fluid, but more importantly, isotope 3H can cause radioactive pollution to the surrounding environment

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  • A method for detecting the immunosuppressive function of regulatory T cells
  • A method for detecting the immunosuppressive function of regulatory T cells
  • A method for detecting the immunosuppressive function of regulatory T cells

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Embodiment Construction

[0027] Below in conjunction with accompanying drawing, the present invention will be further described as follows:

[0028] Method steps of the present invention comprise:

[0029] 8) Purify CD4 + T cells were washed with PBS phosphate buffered saline;

[0030] 9) 10 7 CD4 + T cells were resuspended in 500 μl pre-warmed PBS phosphate buffer;

[0031] 10) Add an equal volume of CFSE hydroxyfluorescein diacetate succinimide lipid, preheat PBS phosphate buffer 5 μM, and the final concentration of CFSE hydroxyfluorescein diacetate succinimide lipid is 2.5 μM;

[0032] 11) Incubate at room temperature for 2 minutes, protected from light;

[0033] 12) Add an equal volume of fetal bovine serum, incubate at room temperature for 1 minute, and avoid light;

[0034] 13) The complete medium was washed 3 times, counted again, and added to a 96-well plate for 5 days;

[0035] 14) Regulatory T cells were added, at day 5 according to CFSE hydroxyfluorescein diacetate succinate

[0036...

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Abstract

The present invention relates to a method for detecting the immunosuppressive function of regulatory T cells, wherein the steps of the method include: purifying CD4+T cells and washing with PBS phosphate buffer; solution; add an equal volume of CFSE hydroxyfluorescein diacetate succinimide lipid, preheat PBS phosphate buffer 5 μM, the final concentration of CFSE hydroxyfluorescein diacetate succinimide lipid is 2.5 μM; incubate at room temperature 2 minutes; add an equal volume of fetal calf serum, incubate at room temperature for 1 minute; wash the complete medium 3 times, recount, add to a 96-well plate for 5 days; beneficial effects of the present invention: safe, and the labeled fluorescence can be maintained for several days Weekly; can track the process of cell division and proliferation in vivo; high correlation; specific tracking operation is simple and reliable; stable, objective results, cost-effective, and high efficiency.

Description

[technical field] [0001] The invention relates to the technical field of a cell detection method, in particular to a method for detecting the immunosuppressive function of regulatory T cells. [Background technique] [0002] Regulatory T cells, which can be differentiated from natural regulatory T cells or other naive T cells, express CD25 molecules and Foxp3, and their differentiation and function must involve the participation of specific cytokines. It plays an important role in maintaining peripheral immune tolerance, preventing the occurrence of autoimmune diseases and limiting chronic inflammatory diseases. Nowadays, the 3H-TdR incorporation method is often used to measure the immunosuppressive function of regulatory T cells. The principle is that T lymphocytes enter cell cycle mitosis after being activated by ConA or PHA. When entering the S phase of the cell cycle, the amount of DNA synthesized by the cells increases significantly. Tritium (3H)-labeled DNA precursor t...

Claims

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Application Information

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IPC IPC(8): G01N21/64C12Q1/02
Inventor 顾鹏
Owner 顾鹏
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