In vitro and In vivo CD8+ T Cell Suppression Assays

Lu Xie; Guihuan Liu; Yanjun Liu; Yuming Yu

doi:10.21769/BioProtoc.4020

In vitro and In vivo CD8⁺ T Cell Suppression Assays

Bio Protoc. 2021 May 20;11(10):e4020. doi: 10.21769/BioProtoc.4020.

Authors

Lu Xie^{1

2}, Guihuan Liu³, Yanjun Liu³, Yuming Yu^{1

2}

Affiliations

¹ Department of Urology, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Guangzhou, China.
² School of Medicine, South China University of Technology, Guangzhou, China.
³ Department of Immunology, School of Basic Medical Science, Southern Medical University, Guangzhou, China.

Abstract

CD8⁺CD28^- T suppressor cells (Ts) have been documented to promote immune tolerance by suppressing effector T cell responses to alloantigens following transplantation. The suppressive function of T cells has been defined as the inhibitory effect of Ts on the proliferation rate of effector T cells. ³H-thymidine is a classical immunological technique for assaying T cell proliferation but this approach has drawbacks such as the inconvenience of working with radioactive materials. Labeling T cells with CFSE allows relatively easy tracking of generations of proliferated cells. In this report, we utilized antigen presenting cells (APCs) and T cells matched for human leukocyte antigen (HLA) class I or class II to study CD8⁺CD28^- T cell suppression generated in vitro by this novel approach of combining allogeneic APCs and γc cytokines. The expanded CD8⁺CD28^- T cells were isolated (purity 95%) and evaluated for their suppressive capacity in mixed lymphocyte reactions using CD4⁺ T cells as responders. Here, we present our adapted protocol for assaying the Ts allospecific suppression of CFSE-labeled responder T cells.

Keywords: CD8+CD28- T cells; Alloantigen specific tolerance; Common gamma chain cytokines; Immune tolerance; Suppression.