Enrichment of normal progenitors in counter-flow centrifugal elutriation (CCE) fractions of fresh chronic myeloid leukemia leukapheresis products.
OBJECTIVE
The aim of this study was to assess the suitability of a technique based on counter-flow centrifugal elutriation (CCE), which should allow one to enrich chronic myeloid leukemia (CML) patients' unstimulated native leukapheresis product (nLP) in CD34+ HLADR- cells and BCR-ABL negative cells.


METHODS
Six newly diagnosed CML patients were subjected to leukapheresis, and the products were subfractionated with the use of CCE. nLP and all fractions were studied for the presence of CD34+ cells and a proportion of BCR-ABL fluorescence in situ hybridization (FISH)+ cells.


RESULTS
CCE fractions with a high flow rate contained the highest proportion of CD34+ cells [mean (SEM) 6.89% (3.88)]. However, CD34+ cells present in low-rate CCE fractions showed a higher proportion of HLADR-[49.6% (13.5 in 70 mL min-1) and 21.5% (11.6 in 110 mL min-1)] than those in 170 mL min-1[3.2% (2.5)] and "rotor off" [3.4% (1.9)]. This was associated with lower proportions of BCR-ABL FISH+[8.1% (4.8) and 1.9 (1.7)] and smaller BCR-ABL to ABL transcript ratios [0.58 (17) and 0.26 (0.08) in 70 and 110 mL min-1] fractions as compared to 140 and 170 mL min-1 fractions [21.6% (5.2) and 31.6% (15.3) for BCR-ABL FISH+ cells and 0.75 (0.16) and 0.90 (0.24) for BCR-ABL/ABL]. Fractions with the lowest proportions of BCR-ABL-positive cells and the lowest BCR-ABL/ABL transcript ratios (110 mL min-1) contained from 1.3 x 106 to 82.7 x 106 (median: 3.97 x 106) CD34+ cells.


CONCLUSIONS
In the present study we have shown that CCE may be used effectively to obtain nLP fractions enriched in normal hematopoietic progenitors.
Dlubek
Dorota
D
Lower Silesian Center for Cellular Transplantation, Institute of Immunology and Experimental Therapy, Polish Academy of Science, Rudolf Weigl 12, 53-114 Wroclaw, Poland.
Dybko
Jaroslaw
J
Wysoczanska
Barbara
B
Laba
Anna
A
Klimczak
Aleksandra
A
Kryczek
Ilona
I
Konopka
Lech
L
Lange
Andrzej
A
D000328:Adult; D018952:Antigens, CD34 / Q000032:analysis; D002469:Cell Separation / Q000379:methods; D002498:Centrifugation / Q000379:methods; D005240:Feasibility Studies; D005260:Female; D016044:Fusion Proteins, bcr-abl / Q000235:genetics; D016313:Genes, abl; D006684:HLA-DR Antigens / Q000032:analysis; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D016130:Immunophenotyping; D017404:In Situ Hybridization, Fluorescence; D007937:Leukapheresis / Q000379:methods* / Q000592:standards; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000097:blood / Q000628:therapy*; D008297:Male; D008875:Middle Aged; D014182:Transplantation, Autologous / Q000379:methods
D018952:Antigens, CD34; D006684:HLA-DR Antigens; D016044:Fusion Proteins, bcr-abl