Although the strain-promoted sydnone-alkyne cycloaddition reaction has been utilized for bioconjugation, its potential applications in bioorthogonal labeling and imaging in live cells have not been explored. This communication reports novel bioorthogonal imaging reagents with environment-sensitive emission properties through the modification of sydnone with cyclometalated iridium(III) polypyridine complexes. These complexes displayed significant emission enhancement and lifetime elongation upon reaction with strained alkyne derivatives, and were utilized to label cyclooctyne-modified proteins and ceramide molecules in live cells. Additionally, the manipulation of the photocytotoxicity of the complexes through the use of a bioorthogonal reagent was demonstrated.