Gradient separation of granulocytic progenitor cells (CFUc) from human blood mononuclear leukocytes

Abstract

Two different density-gradient techniques were compared for separation of human blood-derived granulocyte-macrophage progenitor cells (CFUc) from immunocompetent cells using albumin or Percoll as gradient media. Mean CFUc enrichment by means of discontinuous albumin gradient was twofold, whereas Percoll gradients yielded about 20- to 40-fold enrichment of CFUc in relation to lymphocytes. The pH of gradient media proved to have a major influence on separation quality. Based on these results and the development of a new density-separation technique in a 600-ml plastic bag, replacing customary Ficoll-Hypaque gradients, a method for large-scale purification of CFUc from leukapheresis-derived leukocytes has been constructed, working in a closed, sterile system. About 11 X 10(9) mononuclear cells containing 6 X 10(5) CFUc could be processed within 10 h. Lymphocytes were reduced to less than 1% of their initial number, and an average of about 50% of the original CFUc could be recovered.