Detection of the granulocyte colony-stimulating factor receptor using biotinylated granulocyte colony-stimulating factor: presence of granulocyte colony-stimulating factor receptor on CD34-positive hematopoietic progenitor cells.
Granulocyte colony-stimulating factor (G-CSF) was linked to NHS-biotin to yield biotinylated G-CSF (b-G-CSF), which retained the ability to stimulate colony formation by normal bone marrow (BM) cells in methylcellulose. The use of streptavidin-phycoerythrin conjugate in conjunction with flow cytometry demonstrated that the binding of biotinylated G-CSF to its receptor is saturable, competitive, and specific. A 100-fold molar excess of unlabeled G-CSF almost completely inhibited the binding of the biotinylated G-CSF to the human leukemia cell line U937, which is known to possess the G-CSF receptor. G-CSF receptors were clearly detected by flow cytometry on adult human peripheral granulocytes and monocytes, but not on lymphocytes. Using this method, the expression of G-CSF receptors on hematopoietic progenitor cells in bone marrow and umbilical cord blood, detected as CD34-positive (CD34+) cells, were examined. A small but significant number of CD34+ cells were detected among the bone marrow mononuclear cells and umbilical-cord-blood mononuclear cells (4.28% +/- 0.31%, 1.09% +/- 0.20%, respectively). The percentage of CD34+ BM mononuclear cells was significantly higher than for cord blood mononuclear cells (P less than 0.01). These CD34+ cells were then analyzed by biotinylated G-CSF binding. CD34+ cells from bone marrow contained 25.8% +/- 7.9% G-CSF receptor positive cells and those from cord blood possessed 29.2% +/- 7.0% of G-CSF receptor-positive cells. The difference was not statistically significant.
Shimoda
K
K
First Department of Internal Medicine, Faculty of Medicine, Kyushu University, Fukuoka, Japan.
Okamura
S
S
Harada
N
N
Niho
Y
Y
D015703:Antigens, CD / Q000378:metabolism*; D018952:Antigens, CD34; D001710:Biotin / Q000378:metabolism*; D005434:Flow Cytometry; D016179:Granulocyte Colony-Stimulating Factor / Q000378:metabolism*; D006098:Granulocytes / Q000378:metabolism; D006412:Hematopoietic Stem Cells / Q000378:metabolism*; D006801:Humans; D009000:Monocytes / Q000378:metabolism; D016188:Receptors, Granulocyte Colony-Stimulating Factor / Q000378:metabolism*; D011994:Recombinant Proteins; D014407:Tumor Cells, Cultured
D015703:Antigens, CD; D018952:Antigens, CD34; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor; D001710:Biotin