Comparison of purity and enrichment of CD34+ cells from bone marrow, umbilical cord and peripheral blood (primed for apheresis) using five separation systems.
Interest in the isolation and characterization of primitive hemopoietic cells in both the clinical and research fields has rapidly increased. In parallel, different purification systems have been developed to isolate these cells. We have compared five different methods for separation of CD34+ cells from human umbilical cord blood, normal bone marrow and apheresis harvests and analyzed purity, recovery, yield and enrichment of colony forming cells (CFC) for each individual system. Our results indicate that the most reliable methods of purification for all samples were fluorescence activated cell sorting (FACS) and magnetic activated cell sorting (MACS) which consistently yielded high purities (> 70%) and enrichment of CFC. In this respect the enrichment of CFC from the MACS was superior to all the other systems including FACS. Similar results (> 70%) for purity were obtained using avidin affinity columns and a biotinylated antibody but neither yield nor CFC enrichment approached the values achieved with MACS. On average CFC enrichment using these affinity columns was greater than that observed for FACS while the purity was comparable. Both CELLector flasks and immunomagnetic beads coated with CD34 antibodies were less effective in our hands in separating purified populations of progenitor cells. Both purity and CFC enrichment of CD34+ cells using these methods were at least 50% lower than obtained with either FACS, MACS or affinity columns.
de Wynter
E A
EA
Cancer Research Campaign Department of Experimental Haematology, Paterson Institute for Cancer Research, Manchester, United Kingdom.
Coutinho
L H
LH
Pei
X
X
Marsh
J C
JC
Hows
J
J
Luft
T
T
Testa
N G
NG
D015703:Antigens, CD / Q000032:analysis; D018952:Antigens, CD34 / Q000032:analysis*; D001773:Blood Cells / Q000166:cytology*; D001781:Blood Component Removal; D001854:Bone Marrow Cells; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D046508:Culture Techniques / Q000379:methods; D005260:Female; D005312:Fetal Blood / Q000166:cytology*; D005434:Flow Cytometry / Q000379:methods; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000276:immunology; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods; D007231:Infant, Newborn; D011247:Pregnancy
D015703:Antigens, CD; D018952:Antigens, CD34