Loss of Gfi1b increases the frequency of hematopoietic stem cells (HSC) in bone marrow, spleen, and blood. (A) Course of pIpC treatment of MxCre tg Gfi1b^fl/fl mice and gating strategy to determine HSC and MPP frequencies using the indicated markers to stain bone marrow cells. Loss of Gfi1b significantly enhances the number of HSCs defined as LSK, CD150⁺,CD48⁻. Results are representative for at least 3 independent experiments. (B) Frequency of HSCs in the bone marrow (n = 14) of wt and Gfi1b-deficient mice was determined by flow cytometry (P ≤ .001 for both) 30 days after the first (equivalent to 21 days after the last) pIpC injection. (C) Frequency of CD34⁺ and CD34⁻ HSCs in the bone marrow (n = 4) of wt and Gfi1b-deficient mice was determined by flow cytometry (P ≤ .01) 30 days after the first (equivalent to 21 days after the last) pIpC injection. (D) Frequency of HSCs in the spleen of wt (n = 3) and Gfi1b- (n = 5) deficient mice was determined by flow cytometry (P ≤ .01) 30 days after the first (equivalent to 21 days after the last) pIpC injection. (E) Frequency of HSCs in the peripheral blood (n = 6) of wt and Gfi1b-deficient mice was determined by flow cytometry (P ≤ .01 for both) 30 days after the first (equivalent to 21 days after the last) pIpC injection. (F) Gfi1b^fl/fl and MxCre tg Gfi1b^fl/fl were treated with pIpC, and 30 days after the first injection, peripheral blood cells were analyzed by an Advia blood analyzer. Loss of Gfi1b decreases platelet numbers (n = 6 for Gfi1b^fl/fl and MxCre tg Gfi1b^fl/fl) (P ≤ .01). (G) As in F for red blood cells (n = 6; P ≤ .01). (H) As in F for leukocytes. (I) Genotyping of sorted HSC from pIpC-injected MxCre tg Gfi1b^fl/fl mice. Excision of the Gfi1b allele was efficient, and nonexcised alleles are below detection limit in HSCs.