Isolation of human megakaryocytes by immunomagnetic beads

Br J Haematol. 1989 Sep;73(1):18-22. doi: 10.1111/j.1365-2141.1989.tb00212.x.

Abstract

A simple method was developed to purify human megakaryocytes to homogeneity from normal bone marrow aspirates. An initial separation of marrow between 1.020 and 1.050 g/ml. Percoll density cut was used to enrich megakaryocytes. After washing, the cells were suspended with immunomagnetic beads which were coated with sheep anti-mouse IgG antibody and treated with anti-human glycoprotein (GP) IIb/IIIa monoclonal antibody, or the cells were treated with human platelet GP IIb/IIIa monoclonal antibody and suspended with the immunomagnetic beads which were coated with sheep anti-mouse IgG antibody. Megakaryocytes were selectively separated using a magnet. All of the isolated cells were morphologically recognizable megakaryocytes. 1.5-3.1 x 10(4) megakaryocytes were obtained from 1.7-4.5 x 10(8) bone marrow nuleated cells. These cells were all positive in immunoenzymatic staining for GP IIb/IIIa. Megakaryocytes obtained by this method responded to recombinant human GM-CSF (rhGM-CSF) showing an increased 3H-thymidine (3H-dT) incorporation. These data show that this method is useful for obtaining pure megakaryocyte populations which can be submitted to comprehensive biological studies.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies, Monoclonal
  • Bone Marrow Cells*
  • Cell Separation / methods*
  • Centrifugation, Density Gradient
  • Colony-Stimulating Factors / pharmacology
  • Granulocyte Colony-Stimulating Factor
  • Granulocyte-Macrophage Colony-Stimulating Factor
  • Growth Substances / pharmacology
  • Humans
  • Magnetics
  • Megakaryocytes* / cytology
  • Recombinant Proteins / pharmacology

Substances

  • Antibodies, Monoclonal
  • Colony-Stimulating Factors
  • Growth Substances
  • Recombinant Proteins
  • Granulocyte Colony-Stimulating Factor
  • Granulocyte-Macrophage Colony-Stimulating Factor