NAMS 2002 Workshop - ICOM 2008

isolation of CD34+ cells was achieved using Pluronic-coated membranes. This
was because the Pluronic coating dissolved into the recovery solution at 4oC,
releasing adhered cells from the surfaces of the membranes during permeation
of human serum albumin solution through these membranes. Dextran and/or
bovine serum albumin solutions were also evaluated for use as recovery
solutions after blood permeation. A high recovery ratio of CD34+ cells was
achieved at 4oC in a process using 20% dextran solution through polyurethane
(PU) membranes having carboxylic acid groups. CD34+ (hematopoietic stem)
cells were efficiently recovered (85% recovery ratio) through PU-COOH
membranes in a process using 20 wt% aqueous dextran as the recovery
solution. This indicated that dextran solution was preferable to HSA and BSA
solutions during the recovery process.
Forraz et al. (Stem Cells 22, 100 (2004)) reported that negative-isolated cells,
which depleted umbilical cord blood mononuclear cells from blood cells
expressing mature hematopoietic markers (glycophorin A, CD2, CD3, CD7,
CD16, CD33, CD38, CD45 and CD56), lineage- negative cells, enriched longterm
culture- initiating cells. The lineage-negative cells maintained and expanded
more primitive hematopoietic stem and progenitor cells than CD34+ and CD133+
cells, and expressed higher levels of the cell-adhesion molecule CD162
[expression ratio (ER) = 16.0%] and CD164 (ER = 96.7%) involved in
hematopoietic progenitors forming bone marrow than CD34 (ER = 14.4%) and
CD133 (ER = 7.0%). Therefore, primitive hematopoietic stem and progenitor cells
tend to adhere to polyurethane (PU) membrane surfaces, due to their expression
of these cell-adhesion molecules on their surfaces.
The exact surface marker for primitive hematopoietic stem and progenitor cells
remains unclear at the current time. Isolating such cells by membrane filtration of
umbilical cord or bone marrow is thought to be more effective than magnetic
bead or flow cytometry sorting methods, because cell separation in membrane
filtration is based not only on cell size, but also on the intensity of cell adhesion to
the membrane surface. Of all methods, membrane separation is likely to provide
the most sanitary and simple isolation of primitive hematopoietic stem and
progenitor cells.