Global monitoring of cellular signaling activity is of great importance for the understanding of the regulation of complex signaling networks and the characterization of signaling pathways deregulated in diseases. Tyrosine phosphorylation of intracellular signaling proteins followed by the recognition and binding of Src homology 2 (SH2) domains are key mechanisms in the downstream transmission of many important biological signals. SH2 domains, comprising 120 members in humans, are small modular protein binding domains that recognize tyrosine phosphorylated signaling proteins with high specificity. Based on these binding properties, the large number of naturally occurring and currently available SH2 domains serve as excellent probes for the comprehensive profiling of the cellular state of signaling activity. Here we have described different experimental strategies for global SH2 profiling: high-resolution phosphoproteomic scanning by far-Western Blot analysis and high-throughput profiling by our recently developed oligonucleotide-tagged multiplex assay (OTM) and Rosette assay.