Haematopoietic stem cell content of murine bone marrow, spleen, and blood. Limiting dilution analysis of diffusion chamber cultures

Suspensions of mouse bone marrow cells, spleen cells, and blood leucocytes were cultured in diffusion chambers in dilution series in order to establish the minimum concentrations of haematopoietic stem cells (HSC). The observed frequencies of empty chambers after seven days of culture conformed to the expected frequencies of a null response in a Poisson distribution. The proportions of empty chambers could therefore be used to estimate the concentrations of HSC in the cell suspensions. The following numbers of HSC per 10⁵ cells were found (with 95% confidence limits): Bone marrow: 50 (44–56). Spleen: 3.5 (2.8–4.3). Blood leucocytes: 1.4 (1.2–1.8). The mean (± standard error) HSC-content per femur, spleen, and milliliter blood when pooling cells from three to six donor mice was 8240 ± 600, 7660 ± 490, and 56 ± 6.5 respectively. For comparison, the HSC concentrations were also determined with the spleen colony technique; the ratio between the HSC-concentrations of bone marrow, spleen, and blood determined with the diffusion chamber technique was similar to that determined with the spleen colony technique.