NAMS 2002 Workshop - ICOM 2008

Biomedical and Biotechnology I – 4
Monday July 14, 11:15 AM-11:45 AM, Honolulu/Kahuku
Separation and Purification of Hematopoietic Stem Cells from Human
Blood through Surface-modified Membranes
A. Higuchi (Speaker), Nat. Central Univ. & Nat. Res. Institute for Child Health & Develop., Tokyo,
Japan, higuchi@ncu.edu.tw
Y. Chang, Chung Yuan Christian University, Taoyuan, Taiwan
R. Ruaan, National Central University, Taoyuan, Taiwan
W. Chen, National Central University, Taoyuan, Taiwan
Efficient cell separation is important for the successful isolation and purification of
blood cells, stem cells and specific tissue cells. Techniques such as
centrifugation, affinity column chromatography, and fluorescence activated cell
sorting (FACS), magnetic cell selection, and membrane filtration are typically
employed for cell separation. The centrifugal separation of cells is a typical
method employed to isolate platelets, leukocytes, mononuclear cells, red blood
cells and non-blood cells. Highly purified cellular preparations are obtained using
FACS or a magnetic cell selection system in conjunction with a fluorescentlylabeled
antibody as the cell-surface marker. Cell separation through membrane
filtration was recently reported by several researchers. Leukocyte removal filters
are commercially available cell separation filters. The stem cells that form blood
and immune cells are known as hematopoietic stem cells. Hematopoietic stem
and progenitor cells bear the CD34 cell surface marker. These cells are thought
to be responsible for the reconstitution of hematopoiesis. Therefore, the
transplantation of CD34+ cells is essential in the therapy of patients with acute
myeloid leukemia, myelodysplastic syndromes, chronic myeloid leukemia and
systemic mastocytosis. In a previous investigation (A. Higuchi et al., J. Biomed.
Mater. Res. 68A, 34 (2004)), cell separation from peripheral blood at fixed blood
permeation speeds (1 ml/min) was investigated using surface-modified
polyurethane (PU) membranes with a fixed pore size of 5 mm, carrying different
functional groups. However, optimal conditions for the purification of CD34+ cells
from blood using membrane filtration were still undetermined. In this study, we
prepared several of the membranes, and conducted further experiments on the
separation of CD34+ cells using the membranes.
Cell separation from peripheral blood was investigated using polyurethane (PU)
foam membranes having 5.2 mm pore size and coated with Pluronic F127 or
hyaluronic acid. The permeation ratio of hematopoietic stem cells (CD34+ cells)
and lymphocytes through the membranes was lower than for red blood cells and
platelets. Adhered cells were detached from membrane surfaces using human
serum albumin solution after permeation of blood through the membranes,
allowing isolation of CD34+ cells in the permeate (recovery) solution. High- yield