Lymphocyte isolation from human spleen by counterflow centrifugation employing two different flow chambers on line

Studies on splenic lymphocytes have hitherto been performed on single cell suspensions depleted of phagocytic cells by adherence to plastic or incubation with carbonyl iron. These techniques have the disadvantages of selective cell loss, suboptimal cell purification and cell activation. This paper describes purification of splenic lymphocytes by the use of counterflow centrifugation (CFC). The method was adapted to overcome pelleting of cells in the separation chamber to form a plug at the inlet and impede adequate flow. By combining 2 different separation chambers on line in 1 rotor this problem was overcome. Of all lymphocytes recovered after CFC 88.8 +/- 1.4% were collected in 2 pooled fractions with a purity of greater than or equal to 98% and a cell viability of 95%. After CFC, 80.8 +/- 12.1% of the viable cells loaded were recovered.