Study of the collection and separation of umbilical cord blood for use in hematopoietic progenitor cell transplantation.

Abstract

Human umbilical cord blood has been used as an alternative source of cells for repopulating bone marrow in allogenic bone marrow transplantation in children. The number of transplantations of umbilical cord blood cells is increasing worldwide. Umbilical cord blood was collected from 52 subjects at a single collection at the time of delivery, and separated using the red blood cell sedimentation technique. Nucleated cells harvested from the fraction enriched with white blood cells were used for an assay to detect colony-forming unit granulocyte-macrophage (CFU-GM) derived cell colonies and a flow cytometric analysis of CD34+ cells under variable conditions. The number of hematopoietic progenitor cells that might be reconstituted to bone marrow was estimated. The mean duration time from the beginning of delivery to complete collection of cord blood was 9.9 min (range 5 to approximately 20 min). The mean volume of umbilical cord blood for the 52 collections was 69.1 ml (range 15-135 ml), containing 1.001% CD34+ cells (range 0.21% approximately 2.63%) and 4 x 10(5) cells of CFU-GM derived colonies (range 0.2 x 10(5) approximately 10.0 x 10(5) cells) within 24 h at 4 degrees C after delivery of the infant. There was no contamination by the mother's lymphocytes according to cytogenetic analysis using pYNH24, which is a probe with a variable number of tandem repeat markers. These findings indicated that umbilical cord blood can be easily collected using the syringe method and separated by the red blood cell sedimentation technique using 6% hydroxyethylstarch. Within 24 h at 4 degrees C, hematopoietic progenitor cells were well detected using an assay for CFU-GM derived colonies and were measured by flow cytometric analysis. However, the instability of the number of hematopoietic progenitor cells must be resolved for safe transplantation of hematopoietic progenitor cells as a source of cells for repopulating bone marrow in children.