Bioorthogonal chemical reactions have emerged as useful tools for labeling biomolecules in living systems. Nitrones belong to a relatively new bioorthogonal system that has demonstrated high versatility and applicability. These compounds show high reactivity toward strained cyclooctynes via the strain-promoted alkyne-nitrone cycloaddition (SPANC) reaction. Transition metal complexes containing a nitrone moiety are weakly emissive due to the efficient non-radiative decay caused by the C=N photoisomerization, but exhibit intense and long-lived emission upon reaction with strained alkynes such as (1R,8S,9s)-bicyclo[6.1.0]nonyne (BCN). In this work, three cyclometalated iridium(III) complexes functionalized with two nitrone units have been designed as novel phosphorogenic bioorthogonal reagents for bioimaging and phototherapeutics. Upon reaction with BCN substrates, the complexes are converted to isoxazoline derivatives, rendering the reaction mixture to display significant emission enhancement and lifetime extension. Notably, the complexes showed a higher reaction rate toward a bis-cyclooctyne derivative containing a TEG linker (bis-BCN) compared with its monomeric counterpart. The complexes exhibited high photocytotoxicity in bis-BCN-pretreated HeLa cells, which was attributed to the enhanced 1O2 photosensitization upon elimination of the nitrone-associated quenching pathway. The crosslinking properties of the complexes mean that they are potential candidates for the construction of polymeric materials with intriguing photophysical and biological characteristics.