Use of Leu3a/3b for the accurate determination of CD4 subsets for measurement of intracellular cytokines.

摘要: Background Identification of human T-helper cell subsets is possible by measurement intracellular cytokines after coincubation lymphocytes with phorbol myristate acetate (PMA), calcium ionophore, and brefeldin A for up to 20 h. However, exposure PMA leads internalization membrane CD4 loss resolution the CD4+ cells. Detection CD3+CD8- cells or preselection prior stimulation more cumbersome than direct We report use Leu3a/Leu3b multiclone accurate determination stimulation. Methods Peripheral blood were isolated from healthy normal donors proportion CD3+ / T was determined flow cytometry before incubation PMA, h using a variety anti-CD4 monoclonal antibodies. Results The Leu3a/3b reagent only antibody capable resolving 98% initial events PMA. Conclusions The higher signal-to-noise ratio associated Leu3a3b reagent, compared other CD4-specific antibodies available, allows identification subset even treatment Cytometry 44:148–152, 2001. © 2001 Wiley-Liss, Inc.