A kind of isolation and purification of mouse tumor tissue infiltrating CD4 + cd25 + regulatory t cell approach

Abstract

The invention belongs to the field of cell sorting, in particular to a method for separating and purifying mouse tumor tissue infiltrating CD4 + CD25 + Methods for isolation and purification of regulatory T cells. It specifically includes the following steps: (1) Preparation of single-cell suspension of tumor tissue: Weigh 0.2 g of tumor tissue, cut it into small pieces, put it into a tissue grinder, add 5 mL of tissue homogenate to homogenize at 0°C for 2 min, pass through 200 mesh Filter through a filter, collect the filtrate, centrifuge in a vertical centrifuge at 20°C for 15 minutes at 250 g, discard the supernatant and save the precipitate, resuspend the precipitate with 2 mL of 80% fetal bovine serum sample diluent to form a single-cell suspension, and use the sample diluent to adjust the count The concentration is 1×10 8 cells / mL; (2) Separation of mouse tumor tissue infiltrating lymphocytes; (3) CD4 + CD25 + Immunomagnetic bead sorting of regulatory T cells, the collected cells are CD4 + CD25 + Treg. The method of the present invention uses tumor tissue for the first time to prepare a single cell suspension, and the finally obtained CD4 + CD25 + The purity of Treg reached 86.5%.

Description

technical field [0001] The invention belongs to the field of cell sorting, in particular to a method for separating and purifying mouse tumor tissue infiltrating CD4 + CD25 + Methods for isolation and purification of regulatory T cells. Background technique [0002] CD4 + CD25 + Regulatory T cell (Treg) is a cell population with immunosuppressive function, which can inhibit the immune response of autoreactive T cells, inhibit the activation of traditional T cells, and secrete some immunosuppressive factors. In 1995, Sakaguchi et al. reported for the first time that there was a T cell subset expressing IL-2 receptor α chain (IL2Ra, CD25) in autoimmune tolerance, and this CD4 + CD25 + T cells are named regulatory T cells (Regulatory Tcell, Treg). Subsequent studies have found that Treg is a cell population with immune regulation function, which can secrete IL-10 and TGF-β. It can inhibit the immune response of self-T cells through direct contact between cells or indirect...

AI Technical Summary

Problems solved by technology

However, CD4 infiltrated in tumor tissue + CD25 + There are very few Tregs, and the traditional flow sorting method takes too long and the efficiency is too low

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