Isolation in a single step of a highly enriched murine hematopoietic stem cell population with competitive long-term repopulating ability

Journal Article · Tue Aug 15 00:00:00 EDT 1989 · Blood; (USA)

OSTI ID:5472635

Szilvassy, S J; Lansdorp, P M; Humphries, R K; Eaves, A C; Eaves, C J ^[1]

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1. B. C. Cancer Research Centre, Vancouver (Canada)

A simple procedure is described for the quantitation and enrichment of murine hematopoietic cells with the capacity for long-term repopulation of lymphoid and myeloid tissues in lethally irradiated mice. To ensure detection of the most primitive marrow cells with this potential, we used a competitive assay in which female recipients were injected with male test cells and 1 to 2 x 10(5) compromised female marrow cells with normal short-term repopulating ability, but whose long-term repopulating ability had been reduced by serial transplantation. Primitive hematopoietic cells were purified by flow cytometry and sorting based on their forward and orthogonal light-scattering properties, and Thy-1 and H-2K antigen expression. Enrichment profiles for normal marrow, and marrow of mice injected with 5-fluorouracil (5-FU) four days previously, were established for each of these parameters using an in vitro assay for high proliferative potential, pluripotent colony-forming cells. When all four parameters were gated simultaneously, these clonogenic cells were enriched 100-fold. Both day 9 and day 12 CFU-S were copurified; however, the purity (23%) and enrichment (75-fold) of day 12 CFU-S in the sorted population was greater with 5-FU-treated cells. Five hundred of the sorted 5-FU marrow cells consistently repopulated recipient lymphoid and myeloid tissues (greater than 50% male, 1 to 3 months post-transplant) when co-injected with 1 to 2 x 10(5) compromised female marrow cells, and approximately 100 were sufficient to achieve the same result in 50% of recipients under the same conditions. This relatively simple purification and assay strategy should facilitate further analysis of the heterogeneity and regulation of stem cells that maintain hematopoiesis in vivo.

• OSTI ID: 5472635
• Journal Information: Blood; (USA), Vol. 74:3; ISSN 0006-4971
• Country of Publication: United States
• Language: English

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BONE MARROW CELLS
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CELL PROLIFERATION
ANTIGENS
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