Membrane-based cell affinity chromatography to retrieve viable cells.
A novel scheme for the separation and live recovery of one cell type from a mixture of cells using a cell affinity chromatography (CAC) system is demonstrated. An anti-murine IgG was chemically immobilized to a cellophane support via a carbonyldiimidazole (CDI) link. Murine splenocytes flowed over the support, and B-cells were allowed to attach at a shear rate of 15 s-1. Once loading was terminated, the support was washed at a shear rate of 315 s-1 to remove nonspecifically bound cells. Elution of the B-cells was initiated by the transmembrane diffusion of hydrochloric acid (pH 1), supplied to the side of the membrane opposite the cells. At the same time, a shear flow of normal saline was established on the cell side of the membrane, and cells, freed by acid, were retrieved. Results showed that, on average, 250 cells/mm2 attached to antibody immobilized on cellophane surfaces, at a shear rate of 15 s-1, and that attached cells were successfully displaced by acid supplied to the side of the membrane opposite that holding the cells. On average, at least 60% of the B-cells removed by this elution appeared viable, based on a Trypan Blue dye exclusion assay.
Mandrusov
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Department of Chemical Engineering, Columbia University, New York, New York 10027, USA.
Houng
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A
Klein
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Leonard
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EF
D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology; D002469:Cell Separation / Q000379:methods*; D002470:Cell Survival; D002476:Cellophane; D002846:Chromatography, Affinity / Q000379:methods*; D008567:Membranes, Artificial; D051379:Mice; D013154:Spleen / Q000166:cytology
D008567:Membranes, Artificial; D002476:Cellophane