Phosphoflow analysis of in vivo target inhibition. ALL xenografts were treated with vehicle, ruxolitinib, or rapamycin for 72 hours, spleens were harvested, and cells were gated on CD10⁺/CD19⁺/TSLPR⁺ populations (CRLF2_R xenografts) or CD10⁺/CD19⁺ populations (CRLF2_NR xenografts) and analyzed by phosphoflow cytometry for levels of phosphorylated (p)JAK2, STAT5, S6, and 4EBP1. Data were arcsinh-transformed and are represented as histograms of median fluorescent intensities. Down-regulation and up-regulation of phosphorylation relative to vehicle controls are represented by shift to the left (blue) and right (yellow), respectively, on the horizontal axis per the colorimetric scale (bottom). (A) Histograms of (i) unstimulated and (ii) TSLP-stimulated CRLF2_R xenografts. (B) Histograms of unstimulated CRLF2_NR xenografts.