Gating strategy and surface marker expression of stem and progenitor populations in the BM and lineage cells in the spleen.

After preparing single cell suspensions of (A) unperturbed or (B) perturbed miR-221/222-proficient or of (C) unperturbed or (D) perturbed miR-221/222-deficient BM, stem and progenitor cells were stained for flow cytometry analysis. (E) Single-cell suspension of miR-221/222-proficient or miR-221/222-deficient spleen or blood was prepared, and the major hematopoietic lineage cells were stained for flow cytometry analysis. (F) Surface marker expression of the measured hematopoietic stem, progenitor, and lineage cells. HSC were gated on lin− (B220−CD3−CD4−CD8−CD19−CD11c−CD11b−Gr1−NK1.1−TER119−) c-kit+Sca1+Flk2−CD34−CD150+CD48− cells. MPP1 was gated on lin−c-kit+Sca1+Flk2−CD34+CD150+CD48− cells. MPP2 were gated on lin−c-kit+Sca1+Flk2−CD34+CD150+CD48+ cells. MPP3 were gated on lin−c-kit+Sca1+Flk2−CD34+CD150−CD48+ cells. MPP4 were gated on lin−c-kit+Sca1+Flk2+CD34+CD150−CD48+ cells. CLPs were gated on lin−c-kitloSca1loFlk2+IL7R+ cells. CMPs were gated on lin−c-kit+Sca1−CD34+CD16/32− cells. MEPs were gated on lin−c-kit+Sca1−CD34−CD16/32− cells. GMPs were gated on lin−c-kit+Sca1−CD34+CD16/32+ cells. CD4 T cells were gated as CD3+CD4+; CD8 T cells were gated as CD3+CD8+ cell. B cells were gated as CD4−CD8−B220+CD19+ cell. Myeloid cells were gated as CD4−CD8−B220−CD19−CD11b+Gr1− cells. Granulocytes (Gran.) were gated on CD4−CD8−B220−CD19−CD11b+Gr1+ cells. (G) Antibodies used in the analyses. The numerical data can be found in the FlowRepository.org through FR-FCM-Z6PS accession number. BM, bone marrow; CLP, common lymphoid progenitor; CMP, common myeloid progenitor; GMP, granulocyte-myelocyte progenitor; HSC, hematopoietic stem cell; MEP, megakaryocyte-erythroid progenitor; MPP, multipotent progenitor. (TIF)