Activation‐induced CD 137 is a fast assay for identification and multi‐parameter flow cytometric analysis of alloreactive T cells

Summary Detection and isolation of viable alloreactive T cells at the single‐cell level requires a cell surface marker induced specifically upon T cell receptor activation. In this study, a member of the tumour necrosis factor receptor (TNFR) ‐family, CD 137 (4‐1 BB ) was investigated for its potential to identify the total pool of circulating alloreactive T cells. Optimal conditions for sensitive and specific detection of allogeneic‐induced CD 137 expression on circulating T cells were established. Thereafter, CD 137 + alloreactive T cells were phenotypically and functionally characterized by multi‐parameter flow cytometry. Alloantigen‐induced CD 137 expression identified both alloreactive CD 8 + T cells (mean ± standard error of the mean: 0·21 ± 0·07%) and alloreactive CD 4 + T cells (0·21 ± 0·05%). CD 137 + alloreactive T cells were detected in different T cell subsets, including naive T cells, but were found preferentially in CD 28 + T cells and not in the terminally differentiated T cell subset. Upon allogeneic (re‐)stimulation, the cytokine‐producing as well as proliferative capacity of T cells resided mainly within the CD 137‐expressing fraction. About 10% of the CD 137 + alloreactive T cells produced any combination of interferon ( IFN) ‐γ, interleukin ( IL) ‐2 and TNF ‐α. Polyfunctional alloreactive T cells, defined by multiple cytokine expression, were observed infrequently. In conclusion, activation‐induced CD 137 expression is a fast assay allowing for detection and functional analysis of the total alloreactive T cell compartment at the single‐cell level by multi‐parameter flow cytometry.