In this study we describe three different methods for labeling T lymphocytes with cell trace violet (CTV), in order to track cell division in mouse and human cells, in both the in vitro and in vivo setting. We identified a modified method of CTV labeling that can be applied directly to either conventional or spectral flow cytometry, that maintained lymphocyte viability and function, yet minimized dye spill-over into other fluorochrome channels. Our optimized method for CTV labeling allowed us to identify up to eight cell divisions and the replication index for in vitro-stimulated mouse and human lymphocytes, and the co-expression of T-cell subset markers. Furthermore, the homeostatic trafficking, expansion and division of CTV-labeled congenic donor T cells could be detected using spectral cytometry, in an adoptive T-cell transfer mouse model. Our optimized CTV method can be applied to both in vitro and in vivo settings to examine the behavior and phenotype of activated T cells.
Keywords: cell trace violet; congenic adoptive transfer mouse model; conventional flow cytometry; lymphocyte proliferation; spectral flow cytometry.
© 2024 The Authors. Cytometry Part A published by Wiley Periodicals LLC on behalf of International Society for Advancement of Cytometry.