The reversible phosphorylation of serine, threonine and tyrosine plays an
important role in the biology of trypanosomatids. The identification and quantification of phosphorylation site dynamics will not only assist the elucidation of signaling pathways at the molecular level, but will also facilitate drug discovery through improved candidate selection and mode-of-action studies. Recent advances in the field of mass spectrometry-based phosphoproteomics have made the experimental observation and quantitation of thousands of phosphorylation sites feasible outside of specialist mass spectrometry laboratories. In this chapter, an introduction to phosphoproteomic techniques is presented, the current state of knowledge of the trypanosomatid phosphoproteomes is reviewed, and the potential impact of quantitative phosphoproteomics is discussed.