371e Purification of CD45+ Hematopoietic Cells Directly from Human Bone Marrow and Peripheral Blood Using a Flow-Based P-Selectin

371e Purification of CD45+ Hematopoietic Cells Directly from Human Bone Marrow and Peripheral Blood Using a Flow-Based P-Selectin–Coated Microtube

Srinivas Narasipura¹, Joel Wojciechowski², Brian Duffy¹, Jane L. Liesveld³, and Michael King¹. (1) Biomedical Engineering, University of Rochester, Rochester, NY 14627, (2) CellTraffix inc, Rochester, NY 14623, (3) Leukemia/Blood and Marrow Transplant Program, James P. Wilmot Cancer Center, University of Rochester, 601 Elmwood Ave, Rochester, NY 14642

Recently, a flow-based selectin dependent method for capture and enrichment of specific types of blood cells (CD34+ hematopoietic stem and progenitor cells) from bone marrow and peripheral blood has been described. Using a similar approach of perfusing samples through P-selectin coated microtubes, here we show isolation of CD45+ blood cells directly from human bone marrow and plasma depleted peripheral blood to a high purity (90-97%). We observed that secreted P-selectin glycoprotein ligand-1 (PSGL-1) present in the blood plasma inhibited adherence of leukocytes onto P-selectin immobilized on the microtube surface. Hence an additional step of removing plasma from blood cells either by gravity-driven separation or centrifugation was incorporated prior to sample perfusion in the case of peripheral blood to achieve CD45+ cell purification. The process mimics a ubiquitous mechanism of cell trafficking in the body for recruitment of neutrophils during inflammation. The method is straightforward, rapid, and may represent a practical alternative to CD45+ cell enrichment procedures required for chimerism analysis following allogenic stem cell transplantation.

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