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We report the case of a healthy donor who was mobilized for the purpose of performing an unrelated donor transplantation with subcutaneous injections of rhG-CSF. Because of accidental loss of progenitors, 3 days after completing the first collection, the donor was mobilized again with rhG-CSF, and progenitors were collected. While a similar increase in the pre-apheresis leukocyte count was observed in both procedures, fewer mononuclear cells were mobilized during the second rhG-CSF course, resulting in a poor CD34+ yield. These data suggest that an 8-day interval between commencement of rhG-CSF mobilizations is insufficient to predict an efficient collection of hematopoietic progenitors to ensure engraftment after bone marrow transplantation.

Thirty-four patients diagnosed with breast cancer were included in a prospective study evaluating the bone marrow (BM) CD34+/CD71- cell content, as a predictive parameter of the CD34+ cell mobilization after rG-CSF administration. Analysis of the concentration of medullary CD34+/CD71- cells before priming schedules was significantly related with the collection of CD34+ cells in apheresis day 1 (P = 0.03, r = 0.36), apheresis day 1 + day 2 (P = 0.01, r = 0.42) or the total CD34+ cells collected (P = 0.005, r = 0.47). A BM CD34+/CD71- cell concentration greater than or less than a cut-off value of 30/microl was significantly associated with the yield of CD34+ cells collected by cytapheresis procedures (mean values 3.12 x 10(6)/kg, and 2.19 x 10(6)/kg, respectively, P = 0.013). These results suggest that in breast cancer patients undergoing priming with rG-CSF, steady-state BM CD34+/CD71- measurement is a relevant predictive parameter of CD34+ mobilization.

To evaluate the equivalence of 8G12 conjugated with either phycoerythrin (PE) or fluorescein isothiocyanate (FITC), duplicate fluorochrome-8G12 labelling was carried out in 229 samples. Additionally, 53 samples were simultaneously immunostained with FITC-8G12 and PE-8G12. Significantly higher values (p < 0.001) were observed in PE-CD34+ cells when compared with FITC-CD34+ cells both in duplicate and simultaneous analysis. Our data suggest that the choice of fluorochrome is relevant in the measurement of CD34+ cells with 8G12.

Background: The aim of the present study was to investigate the characteristics of the mobilization of hematopoietic precursor cells CD34+ in peripheral blood following stimulation with recombinant granulocytic colony stimulating factor (G-CSF)., Methods: Fourteen patients (10 males, 4 females: mean age 33 years; range 14-58 years) diagnosed with oncohematologic neoplasms, in complete remission were studied. The patients had not received antineoplastic for at least four weeks prior to inclusion in the study. Recombinant G-CSF (8 micrograms/kg) was administered subcutaneously over a minimum of 4 days. Peripheral blood control were performed prior to administration of G-CSF (day 0), the third (day +3) day, and the sixth day (day +6). Daily leukapheresis was initiated at day +3 in 5 patients and at day +4 in 9 patients. The CD34+ cell content was determined in both peripheral blood and leukapheretic material by flow cytometry with an anti CD-34 monoclonal antibody conjugated with fluorescein., Results: No significant differences were observed between the mononuclear cells and CD34+ counts obtained at the first apheresis and those performed at days +3 or +4 (32 +/- 14 x 10(9) vs 29 +/- 19 x 10(9) and 240 +/- 125 x 10(6) vs 162 +/- 160 x 10(6), respectively). The content of the apheresis products in CD34+ cells correlated positively with the number of these cells circulating in peripheral blood (r = 0.53, p = 0.001). In the second apheresis, the presence of mononuclear cells decreased approximately 20% with respect to the first, remained constant in later collections. To the contrary, a constant maintained decrease was observed in the collection of CD34+ on each leukaphesis in that the fourth apheresis only contributed in approximately 10% of the total quantity of CD34+ cells collected., Conclusions: Maximum mobilization of precursor cells was achieved on the third day at a dosage of 8 micrograms/kg/day, with the data found suggesting that three leukapheretic procedures are enough to collect most of the CD34+ cells mobilized.

Although cryopreservation of human bone marrow has become very common in modern medicine, the knowledge on the effects of this procedure on hemopoietic cells is still limited. We herein have investigated whether the process of concentrating of human bone marrow to its buffy coat, the exposure to the cryoprotectant dimethyl sulfoide (DMSO), and/or the rate of controlled freezing/thawing procedures modifies the flow cytometric analysis of human bone marrow cells. We found that both the exposure of marrow cells to DMSO and/or the freezing procedure significantly modifies both the relative proportions of hemopoietic cell subsets and the intensity of expression of certain surface antigens. Thus, percentages of cells expressing CD7, CD13, CD33 and CD34, were found to be lower in both cryopreserved buffy coat and buffy coat merely exposed to DMSO, in comparison to those in untreated coat samples. Moreover, the intensity of the surface expression of CD33 decreased in cells exposed to DMSO, and further in cryopreserved samples. By contrast, the intensity of the CD19 antigen was higher in the last groups of samples than in the buffy coat or the unfractionated human bone marrow. Our present results suggest that flow-cytometric analysis of cryopreserved human bone marrow cells is not fully equivalent to that corresponding to fresh bone marrow or its fractionated buffy coat.