Experimental Immunology Branch, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, MD 20829, USA.
STAR Protoc. 2020 Dec 1;1(3):100195. doi: 10.1016/j.xpro.2020.100195. eCollection 2020 Dec 18.
Assessing IL-2-induced phosopho-STAT5 (pSTAT5) content can reveal the cytokine responsiveness of individual T cells. Identifying distinct T cell subsets by nuclear transcription factors, such as Foxp3, and concurrently quantifying intracellular pSTAT5, however, has been technically challenging. Conventional Foxp3 staining buffers quench pSTAT5 signals, while commonly used pSTAT5 staining protocols fail to detect Foxp3. The current protocol resolves these issues by describing a procedure to assess IL-2-induced pSTAT5 contents in Foxp3 CD4 Treg cells using multiparameter flow cytometry. For complete details on the use and execution of this protocol, please refer to Waickman et al. (2020).
评估白细胞介素 2 诱导的磷酸化 STAT5（pSTAT5）含量可以揭示个体 T 细胞对细胞因子的反应性。然而，通过核转录因子（如 Foxp3）鉴定不同的 T 细胞亚群，并同时定量细胞内的 pSTAT5，在技术上具有挑战性。传统的 Foxp3 染色缓冲液会淬灭 pSTAT5 信号，而常用的 pSTAT5 染色方案则无法检测 Foxp3。本方案通过描述一种使用多参数流式细胞术评估 Foxp3 CD4 Treg 细胞中白细胞介素 2 诱导的 pSTAT5 含量的程序，解决了这些问题。有关该方案的使用和执行的完整详细信息，请参阅 Waickman 等人（2020 年）。
