title: Optimized processing of growth factor mobilized peripheral blood CD34+ products by counterflow centrifugal elutriation.
abstract: Cell separation by counterflow centrifugal elutriation has been described for the preparation of monocytes for vaccine applications, but its use in other current good manufacturing practice (cGMP) operations has been limited. In this study, growth factor-mobilized peripheral blood progenitor cell products were collected from healthy donors and processed by elutriation using a commercial cell washing device. Fractions were collected for each product as per the manufacturer's instructions or using a modified protocol developed in our laboratory. Each fraction was analyzed for cell count, viability, and blood cell differential. Our data demonstrate that, using standard elutriation procedures, >99% of red blood cells and platelets were removed from apheresis products with high recoveries of total white blood cells and enrichment of CD34+ cells in two of five fractions. With modification of the basic protocol, we were able to collect all of the CD34+ cells in a single fraction. The CD34-enriched fractions were formulated, labeled with a ferromagnetic antibody to CD34, washed using the Elutra device, and transferred directly to a magnetic bead selection device for further purification. CD34+ cell purities from the column were extremely high (98.7 ± 0.9%), and yields were typical for the device (55.7 ± 12.3%). The processes were highly automated and closed from receipt of the apheresis product through formulation of target-enriched cell fractions. Thus, elutriation is a feasible method for the initial manipulations associated with primary blood cell therapy products and supports cGMP and current good tissue practice-compliant cell processing.
authors.affiliation: Beckman Research Institute of the City of Hope, Duarte, California CA 91010, USA.
authors.forename: Chy-Anh
authors.initials: CA
authors.lastname: Tran
authors.forename: Monica
authors.initials: M
authors.lastname: Torres-Coronado
authors.forename: Agnes
authors.initials: A
authors.lastname: Gardner
authors.forename: Angel
authors.initials: A
authors.lastname: Gu
authors.forename: Hieu
authors.initials: H
authors.lastname: Vu
authors.forename: Anitha
authors.initials: A
authors.lastname: Rao
authors.forename: Lan-Feng
authors.initials: LF
authors.lastname: Cao
authors.forename: Amira
authors.initials: A
authors.lastname: Ahmed
authors.forename: David
authors.initials: D
authors.lastname: Digiusto
mesh_terms: D000328:Adult; D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism*; D002454:Cell Differentiation; D002469:Cell Separation / Q000295:instrumentation / Q000379:methods* / Q000592:standards*; D002498:Centrifugation; D006801:Humans; D018189:Immunomagnetic Separation; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology*; D051379:Mice; D016688:Mice, Inbred NOD; D009000:Monocytes / Q000166:cytology*; D010641:Phenotype
chemical_list: D018952:Antigens, CD34; D036341:Intercellular Signaling Peptides and Proteins