Intracellular fluorescent labelling of cells for analysis of lymphocyte migration

A procedure for analysing in vivo migration of lymphocytes labelled in vitro using intracellular fluorochromes is described. Comparison of carboxyfluorescein diacetate, a cytoplasmic label, with Hoechst dye No. 33342 (H33342), a DNA-binding fluorochrome, indicated that H33342 is superior. The concentration of H33342 used for labelling does not significantly affect viability or lymphocyte migration and permits long-term visualization. H33342 allows quantitation of in vivo migration in cell suspension and histological localization in frozen sections. Fluorescence is retained in fixed frozen sections for at least 3 months. This method can be used for analyses of lymphocyte migration and maturation.