IMMUNOMAGNETIC SELECTION OF PURIFIED MONOCYTE AND LYMPHOCYTE POPULATIONS FROM PERIPHERAL-BLOOD MONONUCLEAR-CELLS FOLLOWING CRYOPRESERVATION

Cryopreservation is a method commonly used to store human blood sample s, We sought to determine if cryopreserved peripheral blood mononuclea r cells (PBMC) could be separated effectively into distinct population s by using monoclonal antibodies and immunomagnetic microspheres, PBMC obtained from healthy blood donors and from human immunodeficiency vi rus-infected subjects were cryopreserved for as long as 18 months, Rec overed cells were separated into CD14(+) monocytes and CD4(+) T-cell s ubsets by immunomagnetic selection, Flow cytometry analysis indicated >95% depletion of monocytes from PBMC following immunomagnetic selecti on with anti-CD14, A highly enriched population of CD4(+) T cells was obtained from the CD14-depleted cell fraction by using an anti-CD4 mon oclonal antibody and detachable immunomagnetic beads, The CD4(+) T cel ls were subsequently separated into CD4(+) CD45RO and CD4(+) CD45RA fr actions, Each fraction contained >90% enrichment for the respective su bpopulation and <5% of the reciprocal subpopulation, No significant di fferences in cell surface expression of leukocyte markers, in efficien cy of selection of PBMC subpopulations, or in mitogen-induced prolifer ation were detected in freshly isolated or cryopreserved cells, Effici ent recovery of cryopreserved specimens means that targeted assays can be performed on selected, prospectively stored samples once clinical endpoints have been achieved.