title: Colony forming cell assays for human hematopoietic progenitor cells.
abstract: Hematopoietic stem cells (HSCs) present in small numbers in adult bone marrow (BM), peripheral blood (PB) and umbilical cord blood (CB) produce a heterogeneous pool of progenitors that can be detected in vitro using colony forming cell (CFC) assays. Hematopoietic progenitor cells proliferate and differentiate to produce colonies of maturing cells when cultured in a semisolid methylcellulose-based medium that is supplemented with suitable growth factors and other supplements. The colonies are then classified and enumerated in situ by light microscopy or an automated imaging instrument. CFC assays are important tools in basic hematology research but are also used by clinical cell processing laboratories to measure the progenitor cell content of BM, CB and mobilized PB (MPB) preparations used for cell transplantation. Standard CFC assays for human progenitor cells require a culture period of at least 14 days to enable optimal outgrowth and differentiation of the maximum number of CFCs in a cell preparation. In this chapter protocols are described for the detection and enumeration of myeloid multipotential progenitors and committed progenitors of the erythroid, monocyte, and granulocyte lineages in samples from human PB, MPB, BM, and CB. In addition protocols are described for a modified version of the CFC-assay that allows accurate enumeration of total CFC numbers in CB or MPB after a culture period of only 7 days, but without distinction of colony types.
authors.affiliation: STEMCELL Technologies, Vancouver, BC, Canada. bert.wognum@stemcell.com
authors.forename: Bert
authors.initials: B
authors.lastname: Wognum
authors.forename: Ning
authors.initials: N
authors.lastname: Yuan
authors.forename: Becky
authors.initials: B
authors.lastname: Lai
authors.forename: Cindy L
authors.initials: CL
authors.lastname: Miller
mesh_terms: D000643:Ammonium Chloride / Q000737:chemistry; D002452:Cell Count; D002469:Cell Separation; D003114:Colony-Forming Units Assay / Q000379:methods*; D003470:Culture Media / Q000737:chemistry; D041905:Erythroid Cells / Q000166:cytology; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D008533:Megakaryocytes / Q000166:cytology; D008747:Methylcellulose / Q000737:chemistry; D013535:Suspensions; D013997:Time Factors
chemical_list: D003470:Culture Media; D013535:Suspensions; D000643:Ammonium Chloride; D008747:Methylcellulose