Identification of Hematopoietic Stem Cells by the SE-9000^TM Automated Hematology Analyzer in Peripheral Blood Stem Cell Harvest Samples

Takahisa Yamane, Department of Hematology, Osaka City University Medical School, 1-5-7 Asahi-machi, Abeno-ku, Osaka 545 (Japan) Peripheral blood stem cell (PBSC) transplantation is now used extensively to provide rapid and durable hematopoietic reconstitution following supralethal myeloabla-tive therapies [1]. Although the quantification of the cells responsible for reconstitution is a major clinical issue, there has been no convenient method to identify the presence of stem cells in PBSC harvest (PBSCH) samples, and until now, flow-cytometric identification or stem cell cul-turing has been used for detection. However, these methods require time, technical expertise and expensive reagents. We have already reported a simple and easy method of identifying the stem cells in fresh PBSCH samples using a conventional blood cell counter (SE-9000TM; TOA Medical Electronics, Kobe, Japan) with a white blood cell (WBC) differential function [2]. The SE-9000 measures WBC immaturity using a separate Immature Information (IMI) channel [3]. This report describes our investigation of the correlation between the ratio of CD34+ cells and the ratio of the cells detected by the IMI channel (IMI+ cells) for WBC counts in fresh and frozen PBSCH samples. Between February 1995 and March 1996, 49 PBSCH samples were collected from 48 patients with hematologi-cal malignancies at our hospital using a CS-3000 plus blood cell separator (Baxter Health Care Corporation Fenwal Division, Dearfield, 111., USA). The total blood volume processed in each apheresis was 7 liters. In 16 patients from whom mobilized PBSCs were to be collected, granulocyte-colony-stimulating factor (G-CSF) (Chugai, Tokyo, Japan) was administered at a dose of 5 μg/kg/day, given intravenously 2 days before the start of aphresis. In 32 patients treated by conventional chemotherapy, PBSCs were collected on the day the WBC counts exceeded 5 × 109/1 after the administration of G-CSF (5 μg/kg/day, given intravenously). The product volume was 50 ml. For cryopreservation, hydroxyethyl starch and dimethylsulfoxide were added at concentrations of 6 and 5 %, respectively. The cells were stored at -85°C in a mechanical freezer (Hitachi, Tokyo, Japan). Cryopreserved samples were thawed rapidly in a water bath at 37°C and adjusted to a leukocyte count of 10 × 109/1 using phosphate-buffered saline. The staining antibodies included