Phosphorylation is the addition of a phosphate (PO4) group to a protein or other organic molecule. Phosphorylation activates or deactivates many protein enzymes, causing or preventing the mechanisms of diseases such as cancer and diabetes. This book shows how to use mass spectrometry to determine whether or not a protein has been correctly modified by the addition of a phospate group. The book provides a combination of detailed step-by-step methodology for phosphoproteomic sample preparation, mass spectral instrumental analysis, and data interpretation approaches. It also includes the use of bioinformatic internet tools such as the Blast2GO gene ontology (GO) tool, used to help understand and interpret complex data collected during these studies.