Use of hollow fiber membrane filtration for the removal of DMSO from platelet concentrates

Abstract

It has been hypothesized that, in addition to freezing injury, some damage to platelets may result from the cell packing that occurs during removal of the cryoprotectant. This study examined DMSO removal by fluid exchange across hollow-fiber (HF) filters as an alternative to centrifugation. The DMSO solution with or without cell suspension was passed once through the filter. The optimum exchange during unloading of DMSO was determined by varying the flow rates in the external and internal compartments of the HF filter. Initially, buffered solutions of a 5% DMSO solution in the absence of platelets were pumped into the fibers and exchanged against PBS. The residual DMSO was determined by osmometry. The exchange of DMSO across the membrane was flow dependent and also influenced by the chemical nature of the HF fibers. No protocol using a reasonable rate flow through the fibers removed more than 95% of the DMSO in a single pass. The optimum protocol was achieved with polysynthane fibers with an internal flow rate of approximately 20 ml/min and an external flow rate of 100 ml/min. Subsequently, frozen/thawed platelet concentrates in DMSO were washed using centrifugation and compared to the HF filtration method. Platelet quality was assayed by flowcytometry, cell count, morphology and osmotic stress test. Both filtration and centrifugal washing techniques resulted in comparable morphological scores and numbers of discoid cells. When agents reducing platelet activation were added, platelet quality was improved after washing by either technique. The lower platelet osmotic response with HF filtration than with centrifugation while using activation inhibitors was attributed to the remaining amount of the inhibitors. All other parameters tested were similar. The expression of CD62P was equivalent with both techniques, and centrifugation did not activate platelets more than filtration contrary to what was originally anticipated. In conclusion, platelet quality was comparable after washing by either technique but hollow fiber filtration does remove cryoprotectant more rapidly than does centrifugation.