Large scale purification of human blood CD34(+) cells using a nylon-fiber syringe system and immunomagnetic microspheres.
BACKGROUND
To establish an available, economical technique for the large-scale purification of CD34(+) cells we used a nylon-fiber syringe (NF-S) for depletion of adherent cells and then selected CD 34(+) cells from peripheral blood mobilized by G-CSF, using MAb and magnetic heads.


METHODS
PBSC were mobilized and collected from adult, healthy volunteers. With the effect of concentration of anti-CD34 MAb (9C5) on the purification of CD34(+) cells from 1 2 10(8) NF-S treated cells (NF cells), the recovery and the purity of CD34(+) cells was identical for 5, 10, 20, 40 microg of 9C5. In this study, therefore, the concentration of 9C5 was maintained at >5 microg/10(8) NF cells, with a cellhead ratio of 1:10.


RESULTS
When half to one-third of leukapheresis product containing 121.5 + or - 26.0 2 10(8) mononuclear cells (mean + or - SD; n = 6) was processed for CD34(+) cell purification, 5.26 + or - 3.01 2 10(7) total purified cells were obtained with a purity of CD34(+) cells at 94.9 - 8.5%. The numbers of CD34(+) cells and total progenitor cells recovered in this process were 4.71 + or - 2.33 2 10(7) cells and 145.9 + or - 121.8 2 10(5) cells, respectively. The total recovery of CD34(+) cells was 37.0 + or - 21.0%. The depletion of monocyte by NF-S reduced the 9C5 anti-human CD34 MAb needed for purification of CD34(+) cells to one-third. Two patients were grafted with peripheral blood CD34(+) cells selected by this procedure and achieved a rapid and consistent hematopoiesis.


CONCLUSIONS
Our clinical data show that these cells are capable of rapid reconstitution of hematopoiesis after high-dose chemotherapy. The procedure contains an additional step; however, consistent high purity of CD34(+) cells in the purified population and cost reduction were achieved, both critical issues in the better purging of malignant cells and for a wide clinical application.
Koizumi
K
K
Department of Internal Medicine II, Hokkaido University School of Medicine, Sapporo, Japan.
Sawada
K
K
Sato
N
N
Yamaguchi
M
M
Nishio
M
M
Tarumi
T
T
Takano
H
H
Fukada
Y
Y
Ieko
M
M
Yasukouchi
T
T
Sekiguchi
S
S
Koike
T
T
D000328:Adult; D000911:Antibodies, Monoclonal / Q000378:metabolism; D018952:Antigens, CD34 / Q000096:biosynthesis*; D002448:Cell Adhesion; D002469:Cell Separation / Q000295:instrumentation* / Q000379:methods; D016179:Granulocyte Colony-Stimulating Factor / Q000378:metabolism; D019650:Hematopoietic Stem Cell Mobilization / Q000295:instrumentation*; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods*; D007963:Leukocytes, Mononuclear / Q000166:cytology; D018950:Lipopolysaccharide Receptors / Q000096:biosynthesis; D008297:Male; D008863:Microspheres; D009757:Nylons / Q000737:chemistry; D016896:Treatment Outcome
D000911:Antibodies, Monoclonal; D018952:Antigens, CD34; D018950:Lipopolysaccharide Receptors; D009757:Nylons; D016179:Granulocyte Colony-Stimulating Factor