Yu Zheng, Jinwei Xiao, Tadele Eticha, Dejana Bešlić, Saman Hosseinkhani, Yiran Guan, Guobao Xu
Abstract Electrochemiluminescence (ECL) imaging has progressed from a sensitive-intensity-based detection method to a powerful analytical platform capable of resolving biological and chemical heterogeneity at the single-entity level. This review maps the evolution of ECL imaging along three major axes: (i) spatiotemporal resolution, enabled by advanced luminophores, confined and built-in coreactant pathways, nanozyme catalysis, and 3D emissive-layer control; (ii) digitalization, in which analog luminescence is converted into discrete, statistically robust events for ultrasensitive quantification; and (iii) intelligent analysis, where artificial intelligence enhances denoising, emitter localization, dynamic tracking, multiplexed decoding, and kinetic inference. Mechanistic innovations─such as nanoscale confinement, intramolecular coreactant design, in situ conversion of endogenous metabolites, and asymmetric nanostructures─provide brighter, more localized, and biocompatible emission sources. Parallel advances in materials (metal nanoclusters, quantum dots, perovskites, AIE luminogens, Janus particles), imaging platforms (SECL/PECL modes, super-resolution strategies, microfluidics, smartphone/Raspberry-Pi devices, and large-scale bipolar-electrode arrays), and digital/AI pipelines collectively enable the high-fidelity capture of dynamic processes from single molecules to complex 3D spheroids. Together, these developments establish a coherent spatiotemporal–digital–intelligent framework and position ECL imaging as a versatile, scalable, and excitation-free modality for next-generation biosensing, clinical diagnostics, drug screening, environmental monitoring, and functional material characterization.