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Simultaneous monitoring assay for T-cell receptor stimulation-dependent activation of CD4 and CD8 T cells using inducible markers on the cell surface.
- Source :
-
Biochemical & Biophysical Research Communications . Sep2021, Vol. 571, p53-59. 7p. - Publication Year :
- 2021
-
Abstract
- Isolation of antigen (Ag)-specific T cells is an important step in the investigation of T-cell immunity. Activation-induced markers (AIMs), such as CD154/tumor necrosis factor (TNF)/CD107A/CD134/CD137 enable the sorting of Ag-specific T cells without using human leukocyte antigen (HLA)-multimers. However, optimal conditions suitable for simultaneous detection of both Ag-specific CD4 and CD8 T cells have not been investigated. Here, conditions were optimized to simultaneously detect the maximum number of activated CD4 and CD8 T cells in a TCR-dependent manner. First, the frequency of total pools of AIM-positive cells induced by superantigen, staphylococcal enterotoxin B (SEB), stimulation in various culture conditions was monitored and compared side-by-side. The total amount of AIM-positive CD4 T cells, but not CD8 T cells, was significantly abrogated by addition of brefeldin A. TNF-alpha converting enzyme inhibitor treatment effectively increased the TNF-positive population, without affecting other markers' positivity. AIM-positive CD4 T cells and CD8 T cells were detected at least 3 h after stimulation. Furthermore, examination of the multiple combination of each marker revealed that minimum contribution of CD134 on the total pool of AIM-positive cells at this setting, suggesting the essential and non-essential AIMs to maximize the detected number of AIM-positive cells. Taken together, this optimized method will be a useful tool for the simultaneous monitoring the T-cell receptor stimulation-dependent activation of CD4 and CD8 T cells using inducible markers on the cell surface including Ag-specific T cells. [Display omitted] • CD154/TNF/CD134/CD137 combination boosts the detected the AIM-positive CD4 T cells pool. • CD107A/TNF/CD134/CD137 combination boosts the detected the AIM-positive CD8 T cells pool. • This method simultaneously detects human PBMC AIM-positive CD4 and CD8 T cells during short period after stimulation. • CD154/CD107A/TNF/CD137 combination enable us to detect nearly 100% of AIM-positive CD4 T cells and CD8 T cells. [ABSTRACT FROM AUTHOR]
- Subjects :
- *HLA histocompatibility antigens
*ENZYME inhibitors
Subjects
Details
- Language :
- English
- ISSN :
- 0006291X
- Volume :
- 571
- Database :
- Academic Search Index
- Journal :
- Biochemical & Biophysical Research Communications
- Publication Type :
- Academic Journal
- Accession number :
- 151950378
- Full Text :
- https://doi.org/10.1016/j.bbrc.2021.07.037