Leukemia phospho-STAT5-specific antibody. STAT5 was constitutively tyrosine phosphorylated in a HL cell line L540 (Figure 1a) that was established from the bone marrow of an adult female patient with nodular sclerosis. 2 Then, we used a discovery platform consisting of immunoaffinity profiling coupled to mass spectrometry (PhosphoScan) that identifies large numbers of tyrosine phosphorylated proteins, including active kinases. 3 Whole cell lysates were digested with trypsin, phosphopeptides were immunoprecipitated with phosphotyrosine antibody (pY-100), and resultant immunoprecipitates were analyzed by

LC-MS/MS mass spectrometry. Duplicate runs of LC-MS/MS analysis on an LTQ ion trap mass spectrometer identified 733 phosphotyrosine sites in 554 tyrosine phosphorylated proteins in L540 cells (Supplementary Table 1). These data have been deposited in PhosphoSitePlus (http://www. phosphosite. org). Among tyrosine phosphorylated kinases identified, we observed a large number of Tnk1 phosphopeptides corresponding to multiple tyrosine sites, including Y77, Y235, Y277, Y287 and Y353 (Table 1). However, we did not detect any Tnk1 phosphopeptides by MS analysis in other HL cell lines (data