Ten color flow cytometry reveals CD126 as a marker for mature SP4 and SP8 thymocytes (42.10)

Abstract

Abstract We have developed a 10 color flow cytometric staining protocol for the simultaneous detection and characterization of 21 thymocyte subpopulations. These subpopulations include 17 previously described subsets and 4 new subsets. Importantly, this methodology uses only commercially available monoclonal antibodies and therefore could be easily employed by any investigator with access to a standard 4 laser flow cytometer. Using this novel staining scheme we have analyzed mouse thymocytes at discreet stages of development for expression of multiple cytokine receptors. As previously described, we found IL-7Ralpha to be present on some early subsets, upregulated briefly after beta-selection, downregulated on ISP and DP cells, and then re-expressed after positive selection. IL-4Ralpha was also expressed within the early DN1 and DN2 subsets. However, its expression was downregulated prior to beta-selection (in the DN3a subset) and was re-expressed in the DP cells prior to positive selection. IL-6Ralpha (CD126) showed a dramatically different pattern of expression. It was low or negative on all thymocyte subsets with the exception of the CD24low SP4 and SP8 subsets. The CD24low cells represent the more mature SP populations and therefore CD126 expression appears to be a good marker for the detection of mature thymocytes.