Debulking blood stem cell collections by density gradient centrifugation in a closed-vessel system

Abstract

Blood stem cells collected by apheresis are largely mononuclear cells in nature, so manipulation of blood stem-cell components frequently requires more time and reagents than for a marrow harvest. Reducing the nucleated cell number in mobilized blood stem-cell collections, while preserving hematopoietic progenitor content, would make such manipulations simpler and less costly, but only if debulking procedures were not complex. We evaluated separation of light-density cells and enrichment of CD34+ cells from mobilized peripheral blood stem-cell collections by density gradient centrifugation over buoyant density solution 60 (BDS 60) in a single, closed vessel. Fifteen apheresis products from five normal volunteers and eight cancer patients contained 3.44 (range, 1.19-5.51) x 10(10) nucleated cells. Following processing and washing, there was a median 29% recovery of nucleated cells, 79% recovery of CD34+ cells, 2.49-fold enrichment of CD34+ cells, 0.96-log depletion of CD3+ cells, 0.48-log depletion of CD56+ cells, and 0.72-log depletion of CD19+ cells. Results of processing were affected by the variability in composition of the apheresis products. The enrichment of CD34+ cells varied by donor type, and there was a logarithmic relationship between the preprocessing percentage of CD19+ cells and the log reduction in CD19+ cells. Recovery of cells after thawing and washing was acceptable for processed cells cryopreserved at concentrations over the range of 0.01-1.5 x 10(8)/mL. These results demonstrate a simple method by which an apheresis product of 1-5 x 10(10) cells can be debulked effectively in a single, closed vessel.