Ikaros inhibits growth of AMKL cells transformed with OTT-MAL. (A) The viability of GFP-sorted 6133 cells expressing GFP alone (MIE), full-length Ikaros (IK1), or the IK7 dominant-negative isoform was assessed over time by trypan blue exclusion. Means ± SD of triplicate experiments are shown. (B) c-Kit, CD41, and CD42 surface marker expression 48 hours after transduction of 6133 cells with MIE, IK1, or IK7. (Left panels) Representative experiment. (Right panels) Bar graph of the means ± SD of the percentages of CD41⁺CD42⁺ cells (n = 3). *P = .0004. (C) Annexin V, 7-AAD staining of 6133 cells was evaluated 24 and 48 hours after transduction by flow cytometry. (Top panel) Representative FACS plot at 48 hours. (Bottom panel) Bar graph depicts means ± SD of triplicate experiments. *P < .05. (D) Cell-cycle analysis of GFP- or IK1-expressing 6133 cells was measured by BrdU incorporation and 7-AAD staining coupled with flow cytometry. *P = .0004. (E) Expression of the Notch target gene Hes1, Nrarp, Kit, Cdkn1a, Dtx2, and Hes6 in 6133 cells expressing GFP or IK1 was measured by qRT-PCR. Means ± SD of 3 independent experiments are shown. *P ≤ .01. 7-AAD, 7-amino actinomycin D; FACS, fluorescence-activated cell sorter.