Open Access

Rapid flow cytometry-based assay for the evaluation of γδ T cell-mediated cytotoxicity

  • Authors:
    • Qili Jin
    • Lina Jiang
    • Qiao Chen
    • Xiaoxiao Li
    • Yinyin Xu
    • Xueqian Sun
    • Ziyue Zhao
    • Li Wei
  • View Affiliations

  • Published online on: December 15, 2017     https://doi.org/10.3892/mmr.2017.8281
  • Pages: 3555-3562
  • Copyright: © Jin et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )
Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )


Abstract

The effector function of natural killer, lymphokine-­activated killer cells and T lymphocytes is commonly evaluated by radioactive chromium‑release cytotoxicity assays. In addition to this indirect method, fluorescence assays have been described for the assessment of in vitro cell‑mediated cytotoxicity. In the present study, target cells were stained with 5‑(and‑6)‑carboxyfluorescein diacetate succinimidyl ester (CFSE), which is a stable integrated fluorescent probe that allows target and effector cells to be distinguished from one another. Staining of target THP‑1 cells with 8 µM CFSE revealed high and stable loading of fluorescence and no effect of the viability of cells. After 4 h of in vitro co‑culture between γδ T cells and CFSE‑labeled infected or uninfected THP‑1 cells, staining with propidium iodide (PI) was performed to distinguish between vital and dead cells. During sample acquisition, target cells were gated on the CFSE positivity and examined for cell death based on the uptake of PI. CFSE and PI double positive cells were recognized as the dead target cells. The percentage of cytotoxicity in the CFSE‑gated cell population was calculated by subtracting the value obtained for non‑specific PI‑positive target cells, which was measured in a control group that did not contain effector cells. The present study describes a simple and convenient assay that is based on the direct quantitative and qualitative analysis of cell damage at a single cell level utilizing a two‑color flow cytometric assay. In conclusion, the flow cytometric‑based assay described in the current study is a simple, sensitive and reliable tool to determine the cytolytic activity of γδ T lymphocytes against mycobacteria. Therefore, the present study may provide valuable information concerning the methods employed to investigate the function of γδ T cells and potentially other lymphocyte subsets.
View Figures
View References

Related Articles

Journal Cover

March-2018
Volume 17 Issue 3

Print ISSN: 1791-2997
Online ISSN:1791-3004

Sign up for eToc alerts

Recommend to Library

Copy and paste a formatted citation
x
Spandidos Publications style
Jin Q, Jiang L, Chen Q, Li X, Xu Y, Sun X, Zhao Z and Wei L: Rapid flow cytometry-based assay for the evaluation of γδ T cell-mediated cytotoxicity. Mol Med Rep 17: 3555-3562, 2018
APA
Jin, Q., Jiang, L., Chen, Q., Li, X., Xu, Y., Sun, X. ... Wei, L. (2018). Rapid flow cytometry-based assay for the evaluation of γδ T cell-mediated cytotoxicity. Molecular Medicine Reports, 17, 3555-3562. https://doi.org/10.3892/mmr.2017.8281
MLA
Jin, Q., Jiang, L., Chen, Q., Li, X., Xu, Y., Sun, X., Zhao, Z., Wei, L."Rapid flow cytometry-based assay for the evaluation of γδ T cell-mediated cytotoxicity". Molecular Medicine Reports 17.3 (2018): 3555-3562.
Chicago
Jin, Q., Jiang, L., Chen, Q., Li, X., Xu, Y., Sun, X., Zhao, Z., Wei, L."Rapid flow cytometry-based assay for the evaluation of γδ T cell-mediated cytotoxicity". Molecular Medicine Reports 17, no. 3 (2018): 3555-3562. https://doi.org/10.3892/mmr.2017.8281