 Human lymphocytes can be isolated most readily from peripheral blood by density centrifugation with the carbohydrate polymer Ficoll Ficoll Histopaque, Sigma, Cat 1077-1. This yields a population of mononuclear cells at the interface that has been depleted of red blood cells and most polymorphonuclear leukocytes or granulocytes. The resulting population, called peripheral blood mononuclear cells, consists mainly of lymphocytes and monocytes. Diluted anticoagulated blood is layered over Ficoll and centrifuges at 400g, 30min at 20oC slow acceleration, no brake. Red blood cells, polymorphonuclear leukocytes or granulocytes have higher density from Ficoll and are to the bottom of the tube. But mononuclear cells consisting of lymphocytes together with some monocytes band over it and can be recovered at the interface. Cells are washed using 1xPBS with 1mM EDTA 3 times to remove Ficoll and platelets. 