Abstract: Bioorthogonal chemistry enables researchers to study biomolecules in their native environments without perturbing endogenous cellular processes. Over the past two decades, significant strides have been made in developing new, and refining existing, bioorthogonal reactions. The continuously expanding toolbox has opened avenues for tackling increasingly complex biological questions. Despite these advances, limitations remain. The majority of bioorthogonal chemistries follow similar mechanisms, and thus cannot be used simultaneously in multicomponent labeling applications. Bioorthogonal chemistry has also seen little use outside of biomolecule labeling and pull-down applications. To push bioorthogonal chemistry into new directions, I took advantage of two l scaffolds previously developed in the lab: the 1,2,4-triazine and the cyclopropenone (CpO). In one project, I leveraged the biocompatibility of CpOs to develop a mild, organocatalytic methodology for synthesizing substituted butenolides. In another, I aimed to exploit the unique reactivity of CpOs with bioorthogonal phosphines to capture protein-protein interactions. Last, I optimized 1,2,4-triazines to develop mutually orthogonal bioorthogonal cycloadditions for multicomponent labeling applications.