The glycome is dynamically regulated during development and disease progression. The ability to detect changes in protein glycosylation, in tissue and serum samples as well as in vivo, is of considerable interest for biomarker identification and disease monitoring. This presentation will focus on our work toward profiling glycomic changes using proteomics and in vivo imaging techniques. Bioorthogonal chemistry is central to our approaches, which involve metabolic labeling of glycans with bioorthogonal functional groups followed by chemical tagging with affinity reagents, mass spectrometry tags or imaging probes. Fundamental principles of bioorthogonal reaction development will be discussed, along with applications in cancer biomarker discovery and glycan imaging during embryonic development.