Aberrant hematopoietic cell development in Ptpn11^D61G/+ mice. (A-B) BM cells freshly harvested from femurs and tibias (both hind limbs) from 5- to 7-month-old Ptpn11^D61G/+ and WT littermates (n = 10 per group) were assayed by multiparameter flow cytometric analysis to determine frequencies of hematopoietic cell populations of various stages and lineages. The cells were stained with antibodies against lineage markers, c-Kit, Sca-1, CD34, Flk2, CD16/32, and IL-7R (CD127). The proportion of BM cells corresponding to the HSC-containing LSK (Lineage⁻Sca-1⁺c-Kit⁺) cell population was quantified. (C-D) Frequencies of LSK cells in spleens of Ptpn11^D61G/+ (n = 5) and WT (n = 4) littermates were determined as above. (E) BM LSK cells were sub-fractioned according to CD34 and Flk2 expression to yield phenotypic assessments of LT-HSC (LSK, Flk2⁻CD34⁻) and ST-HSC (LSK, Flk2⁻CD34⁺) fractions. (G) More differentiated BM LK cell (Lineage⁻c-Kit⁺Sca-1⁻) population was sub-fractioned based on CD16/32 and CD34 expression to identify CMP, GMP, and MEP progenitors (left panel). CLP progenitors were identified by gating L⁻K^lowS^low (Lineage⁻c-Kit ^low Sca-1 ^low) cells followed by assessing CD127 expression (right panel). (F,H) Frequencies of LT-HSC, ST-HSC, CMP, GMP, MEP, and CLP populations in BM cells were quantified (n = 10 per group).