Label 10E6 cellule/ml in 5 µM CFSE final (conc stock 1000x ).
I label the cell at RT for 8 min and block immediately the reaction with FCS (2% final); spin immediately with PBS FCS 2%. Wash 2x after labeling, then you're ready to stimulate.
You have to wait at least one day in culture or in vivo before to be able to find the Facs settings.
Do not let the labeled cells too much time under the hood without any light protection or you will loose the sharpness of the staining.
Sebastien Calbo, Ph.D.