Amplification and identification of regulatory T cells derived from mice in vitro / 中国医师杂志

ABSTRACT

Objective@#To establish a method for in vitro expansion of regulatory T cells (Tregs) for clinical study and immunotherapy.@*Methods@#CD4+ T cells were isolated from BALB/c spleens by magnetic activated cell sorting (MACS), then cultured in 24-well plates coated with anti-CD3/CD28 microbeads in the presence of 100 U/ml interleukin-2 (IL-2) and 5 ng/ml recombinant human transforming growth factor-β (rhTGF-β). The purity of the expanded Tregs were tested by flow cytometry. In vitro inhibition of CD4+ CD25-T cells response by expanded Tregs was measured by mixed lymphocyte reaction test. The number and the viability of the expanded Tregs were detemined by trypan blue staining.@*Results@#Tregs were expanded up to 20 folds after 5 days in culture, and the activity was (93±4)%. The purity of expanded Tregs were (79.1±1.5)%, and maintained suppressive ability.@*Conclusions@#We can obtain large numbers of Tregs with highly purity and suppressive ability, thereby providing a solution to the availability of sufficient Tregs in clinical study and immunotherapy.