Scl expression in HSCs correlates with quiescence. (A) Schematic diagram of hematopoietic populations and their cell-surface markers. HSCs are KSL, which include LT-HSCs (CD150⁺CD48⁻) and ST-HSCs. Progenitors that are devoid of stem cell activity are Kit⁺Sca1⁻Lin⁻, except for CLPs, which are Kit^loSca1^loLin⁻ and express IL7R. Progenitors can be further differentiated on the basis of CD34 and FcγR expression. (B) The cell-cycle status of HSCs and progenitors assessed by Hoechst 33342 (DNA) and Pyronin Y (RNA) staining. G₀, G₁, and S-G₂-M are defined as shown (dot plots are representative of 5 independent experiments). (C) Scl mRNA levels in purified populations from adult mice were assessed by quantitative RT-PCR and normalized using Hprt. Expression levels in LT-HSCs were set as 1 (mean ± SD of 3 experiments). (D) Scl gene expression was monitored by β-galactosidase staining (FDG) and flow cytometric analysis in LT-HSCs, ST-HSCs, and progenitor populations from Scl^+/− mice in which the LacZ gene was inserted into the Scl locus (shaded histograms) and wild-type (white histograms) mice. The mean fluorescence intensities (MFIs) are indicated for Scl^+/− populations. Data shown are representative of 2 independent experiments with groups of 3 or 5 mice each. (E) LT-HSC–enriched populations were further purified into G₀ or G₁/S/G₂/M fractions by flow cytometry with Hoechst and Pyronin Y, as in panel B. Scl and Cdkn1a expression levels were assessed by quantitative RT-PCR. mRNA levels were normalized using Hprt and compared with expression levels in the G₀ population (mean ± SD of 4 replicates; 2 experiments).