Reaction-enabled, highly sensitive super-resolution imaging

ABSTRACT Super-resolution microscopy breaking the optical diffraction limit has led to success in illuminating the complexity of biology with fluorescence—light-excited luminescence 1,2 . Here, we propose another approach for super-resolution imaging with electrochemiluminescence (ECL)—reaction-excited luminescence 3,4 . While featuring high sensitivity in molecular analytics owing to its laser-free excitation, the current reaction-driven imaging suffers from limited resolution due to low photon budget 4 . Efficient ECL imaging of intracellular organelles remains elusive. To meet this challenge, we develop RIED (Reaction-based super-resolution Imaging via Entropy-weighted correlation combined with Deconvolution), which builds upon two pivotal advancements: (i) unprecedented ECL intracellular organelles imaging through a 1,000-fold enhancement in ECL emission, and (ii) the super-resolution ECL imaging by the spatiotemporal acquisition and efficient utilization of highly dynamic ECL profiles. RIED allows for the first three-dimensional super-resolution ECL imaging with a spatial resolution of ∽97 nm in the lateral dimension and ∽235 nm in the axial dimension. This allows to reveal outer mitochondrial membrane topology and distinct microtubule organizations in different mitosis phases, offering a chemistry-based imaging capability for high-throughput imaging and highly sensitive interfacial imaging, which provides a new dimension into biology.