LYMPHOCYTE FRACTIONATION USING IMMUNOMAGNETIC COLLOID AND A DIPOLE MAGNET FLOW CELL SORTER

The relationship between cell function and surface marker expression i s a subject of active investigation in biology and medicine. These inv estigations require separating cells of a homogeneous subset into mult iple fractions of varying marker expression. We have developed a novel cell sorter, the dipole magnet flow sorter (DMFS), which separates se lected T lymphocyte subpopulations, targeted by immunomagnetic colloid , into multiple fractions according to cell surface marker expression, as determined by flow cytometry. A narrow stream of cells is introduc ed into a sheath of carrier fluid in a rectangular channel while subje cted to a perpendicular magnetic force. The special design of the pole pieces ensures a constant magnetic force acting on the magnetically l abeled cells in the separation area. Cells are spread across the flow in relation to their magnetophoretic mobility. Separation is achieved by control of the positions of the effluent stream boundaries, which s eparate fluid volumes with cells of different magnetophoretic mobility . CD4 and CD8 T lymphocytes labeled with primary antibody-fluorescein isothiocyanate (FITC) conjugate and anti-FITC-magnetic colloid are the chosen cell systems. Flow cytometry analysis shows that, for CD4 cell s, a three-fold increase in total marker number per cell is observed w hen comparing the highest to the lowest fluorescence fractions. Simila rly, a four-fold increase in total marker number is observed for CD8 c ells. We also observed the separation of two dissimilar cell types tha t differed in expression of the CD4 marker, monocytes and T helper lym phocytes. We believe that this type of separation is applicable to any cells in suspension for which a suitable antibody exists and, due to the comparatively gentle nature of the process, is particularly suitab le for the sorting of fragile cells. (C) 1998 Elsevier Science B.V. Al l rights reserved.