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Isolation of human thymocytes differing in maturation state and function by centrifugal elutriation.

A modified centrifugal elutriation technique was used to separate (up to 3 X 10(9)) human thymocytes, according to size in 6 different fractions. Eighty percent of the unfractionated thymocytes were recovered in fractions 1 and 2. The majority of these thymocytes appeared to be small and phenotypically immature as was determined by the high percentage of cells reacting with the monoclonal antibodies OKT-6, Mas-036 and peanut agglutinin. In addition, a relatively low percentage of the cells reacted with a monoclonal antibody directed against HLA-A, B and C determinants (Mas-015). The immaturity of these thymocytes was confirmed by their failure to respond to phytohemagglutinin (PHA) and their negligible responder capacity in mixed leukocyte cultures. Fractions 3-6, representing 20% of the unfractionated thymocytes, were collected arbitrarily and contained thymocytes of various maturation stages as judged by their phenotype. The PHA responsiveness and responder capacity in mixed leukocyte cultures of the thymocytes in these fractions were, in general, considerably higher than those of the unfractionated thymocytes. Our data indicate that centrifugal elutriation is a fast and reproducible method to separate large quantities of functionally inactive and phenotypically immature thymocytes from the more mature and functionally active thymocytes.