IMMUNOLOGICAL RESPONSIVENESS OF FROZEN-THAWED HUMAN LYMPHOCYTES.

Mononuclear cells (10-20 × 106) obtained from human peripheral blood by a standard Ficoll-Hypaque technique were suspended in RPMI 1640 media at 4°C containing 10% foetal calf serum and 7.5% dimethyl sulphoxide (DMSO). Two- millilitre aliquots were cooled at - 1°C/min in a Cryoson BV-4 programmed freezing system to - 30°C, then - 5°C/mm to - 80°C and stored in liquid nitrogen vapor. On the day of testing, cell suspensions were thawed rapidly in a 37°C water bath. DMSO was diluted slowly out of the sample and cells resuspended in fresh RPM [1640. it was found that frozen stored human lymphocytes (FSHL) demonstrated all the characteristics of fresh unfrozen cells. These included their ability to form spontaneous rosettes with sheep erythrocytes (`E' rosettes) and sheep erythrocyte-antibody--complement rosettes (`EAC' rosettes). The presence of surface immunoglobulins and Fc receptors were shown by membrane immunofluorescence to be comparable. in addition, the results show that FSHL respond to mitogens, specific antigens; act as both stimulators and responders in the mixed lymphocyte culture reaction; and exhibit cell-mediated lymphocytotoxicity following in vitro sensitization, or against antibody-coated target cells. [ABSTRACT FROM AUTHOR]