Isolation of Megakaryocytes Using Magnetic Cell Separation

Abstract

Megakaryocytes are difficult to isolate because of their fragility, their tendency to aggregate, and their varying sizes. An immunomagnetic cell sorting method (MACS) has been developed for isolation of intact human megakaryocytes from whole bone marrow with a high purification and high recovery yield. Human megakaryocytes were isolated using an antibody to platelet GPIIb/IIIa and a second antibody conjugated to magnetic beads. Megakaryocytes averaged 0.05±0.01% of all nucleated cells in starting marrow cell suspensions. An enrichment up to 43.1% of GPIIb/IIIa positive cells was obtained using the MACS technique. Microscopic examination confirmed that most isolated cells were megakaryocytes, most of which were large and had a well preserved structure. Megakaryocytic viability was estimated to be greater than 90% in all samples. The modal ploidy class of the 8 samples examined was 16N and showed the normal distribution of a megakaryocyte population, revealing that both small and large megakaryocytes were recovered by the MACS separation. Immunomagnetic selection of megakaryocytes is therefore a useful method to separate human megakaryocytes, including low ploidy immature megakaryocytes from routine marrow aspirates.