Purification and quantification of T and B lymphocytes by an affinity method.

Abstract

Affinity surfaces were produced by coupling human immunoglobulin (HGG) to the surface of tissue culture grade plastic-ware with a water-soluble carbodiimide followed by treatment with anti-HGG antisera. Surface immunoglobulin SIg) bearing human B lymphocytes attach to these surfaces when centrifuged on to them and unattached cells could be recovered by inverting the trays or dishes. Optimal cell attachment conditions could be rapidly evaluated by counting cells attached to representative areas of multi-well trays and percentage of SIg-bearing cells quantified. Evidence was obtained for cell attachment through Fc receptors as well as SIg using unrelated antigen--antibody-coated trays. This could be prevented by using the F (ab')2 fragments of the antisera. Under these conditions specific attachment through K and lambda light chains could be achieved with normal and chronic lymphocytic leukaemic lymphocytes. Using tissue culture plastic Petri dishes and relatively small quantities of antiserum, larger numbers of lymphocytes could be processed to produce T lymphocytes containing less than 1 per cent of contaminating SIg-positive cells.