title: Current practices and prospects for standardization of the hematopoietic colony-forming unit assay: a report by the cellular therapy team of the Biomedical Excellence for Safer Transfusion (BEST) Collaborative.
abstract: OBJECTIVE
  Wide acceptance of the colony-forming unit (CFU) assay as a reliable potency test for stem cell products is hindered by poor inter-laboratory reproducibility. The goal of this study was to ascertain current laboratory practices for performing the CFU assay with an eye towards identifying practices that could be standardized to improve overall reproducibility.
  
  
  METHODS
  A survey to evaluate current laboratory practices for performing CFU assays was designed and internationally distributed.
  
  
  RESULTS
  There were 105 respondents to the survey, of whom 68% performed CFU assays. Most survey recipients specified that an automated rather than a manual cell count was performed on pre-diluted aliquots of stem cell products. Viability testing methods employed various stains, and when multiple sites used the same viability stain, the methods differed. Cell phenotype used to prepare working cell suspensions for inoculating the CFU assay differed among sites. Most respondents scored CFU assays at 14-16 days of incubation, but culture plates were read with various microscopes. Of 57 respondents, 42% had not performed a validation study or established assay linearity. Only 63% of laboratories had criteria for determining if a plate was overgrown with colonies.
  
  
  CONCLUSIONS
  Survey results revealed inconsistent inter-laboratory practices for performing the CFU assay. The relatively low number of centers with validated CFU assays raises concerns about assay accuracy and emphasizes a need to establish central standards. The survey results shed light on numerous steps of the methodology that could be targeted for standardization across laboratories.
authors.affiliation: NHS Blood and Transplant North Bristol Park, Filton, UK.
authors.forename: Derwood
authors.initials: D
authors.lastname: Pamphilon
authors.forename: Eileen
authors.initials: E
authors.lastname: Selogie
authors.forename: David
authors.initials: D
authors.lastname: McKenna
authors.forename: Jose A
authors.initials: JA
authors.lastname: Cancelas-Peres
authors.forename: Zbigniew M
authors.initials: ZM
authors.lastname: Szczepiorkowski
authors.forename: Ron
authors.initials: R
authors.lastname: Sacher
authors.forename: John
authors.initials: J
authors.lastname: McMannis
authors.forename: Hermann
authors.initials: H
authors.lastname: Eichler
authors.forename: Henk
authors.initials: H
authors.lastname: Garritsen
authors.forename: Minoko
authors.initials: M
authors.lastname: Takanashi
authors.forename: Leo
authors.initials: L
authors.lastname: van de Watering
authors.forename: David
authors.initials: D
authors.lastname: Stroncek
authors.forename: Jo-Anna
authors.initials: JA
authors.lastname: Reems
mesh_terms: D018952:Antigens, CD34 / Q000378:metabolism; D064987:Cell- and Tissue-Based Therapy; D003114:Colony-Forming Units Assay / Q000592:standards*; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells; D006801:Humans; D012015:Reference Standards; D015203:Reproducibility of Results; D013234:Stem Cells / Q000166:cytology*
chemical_list: D018952:Antigens, CD34