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. 2009 May 28;114(6):1186–1195. doi: 10.1182/blood-2008-09-179788

Figure 3.

Figure 3

Increased cell cycling of myeloid progenitors in Id1−/− mice. (A) Mice received an intraperitoneal injection of 3 mg BrdU and then 1 mg/mL BrdU in the drinking water for 2 days. BMC were isolated from treated mice, depleted of lineage-positive cells, and then stained with IL7Rα, c-Kit, and Sca-1 antibodies to detect LSK and LS−K progenitors. BrdU incorporation was measured by use of the APC BrdU flow cytometry kit (Becton Dickinson). Data are presented as the mean ± SEM of 3 mice for each genotype (*P = .002). (B) Cell proliferation was measured by sorting the indicated number of LS−K or LSK cells from Id1−/− and Id1+/+ mice into 96-well plates (24 replicates per cell concentration) with cIMDM media containing cytokines (mouse SCF, 100 ng/mL; human IL-6, 50 ng/mL; human Flt3, 100 ng/mL; and human Tpo, 100 ng/mL for LSK and mouse SCF,100 ng/mL; mouse IL-3, 30 ng/mL; and mouse GM-CSF, 100 ng/mL for LS−K). Cell proliferation, including colonies (> 50 cells) and clusters (> 10 and < 50 cells), was determined after 10 days of culture, and then the log of the percentage of negative wells was plotted against the number of cells per well. The frequency of a positive colony or cluster is defined as the inverse of the number of seeded cells that corresponds to 37% negative cells.