The integration of bioorthogonal click chemistry with live-cell fluorescent labeling is of paramount significance for elucidating the spatiotemporal information of biomolecules, which is fundamental to understanding core biological processes and disease pathogenesis. Photoclick chemistry has garnered considerable interest as it circumvents the limitations of metal toxicity and lack of spatiotemporal precision inherent in traditional click reactions. This project addresses the critical challenges of conventional photoclick systems, namely the phototoxicity and poor penetration of UV light, alongside the cytotoxicity of substrates activated by visible light. We conducted a comprehensive investigation, proposing novel reaction mechanisms based on tetrazole compounds and strain-promoted cycloadditions with bicyclo[1.1.0]butanes. By leveraging principles from bioorthogonal chemistry, organic synthesis, and molecular biology, we have translated the concept of spatiotemporally controlled live-cell labeling into the development of new cycloaddition reactions and the synthesis of [3.1.1]propellane scaffolds as saturated aromatic ring bioisosteres. Consequently, we have established a series of novel analytical methods for live-cell fluorescence labeling based on this advanced photoclick chemistry. This work not only opens a new avenue for studying intracellular macromolecules via chemical reactions but also significantly expands the repertoire of bioorthogonal transformations. It thereby fosters a deeper integration of chemistry and life sciences, offering promising new strategies and methodologies for applications in bioimaging and therapeutic monitoring.