Relative Notch1, Notch2, Notch3, and Notch4 expression in human hematopoietic marrow progenitor subsets as compared with unfractionated marrow (uF) as measured by real-time PCR. Normal marrow cells were suspended in Hanks solution with 5% human serum at 5 to 10 × 10⁶/mL and incubated for 30 minutes with CD34–fluorescein isothiocyanate (CD34-FITC) (gift from Dr P. Lansdorp) used at 4 μg/mL and CD38-PE (Becton Dickinson) used according to the manufacturer's instructions. Separate aliquots were stained with an irrelevant mouse immunoglobulin G1–FITC (IgG1-FITC) and IgG1-PE antibody (both from Becton Dickinson) as an isotype control. Viable CD34⁺CD38^–, CD34⁺CD38⁺, and CD34^– cells were sorted and used for RNA extraction. The cDNA of unsorted marrow and of marrow sorted for CD34 and CD38 was used for real-time PCR with Notch primers and SybrGreen. Notch expression in the sorted subpopulations was compared with its unfractionated population (uF) to allow comparisons and relative expression of Notch1 through Notch4 in the progenitors expressed. Error bars indicate the standard error of the mean. The expression of the sorted population was compared with the unsorted population in a paired t test. *P ≤ .05. Notch1 is indicated by ▪; Notch2, by □; Notch3, by ▧; and Notch4, by ▦.