When scientists look at cells under a fluorescence microscope, they often see a "fog" or blurry background that hides the real structures they want to study. This happens because the microscope captur...

Mitochondria are excitable organelles, and their electrophysiological activity across the inner mitochondrial membrane (IMM) plays a critical role in energy metabolism, cell signaling and cell fate regulation. While the transmembrane voltage of the IMM (Δψm) is traditionally measured with Nernstian dyes such as tetramethylrhodamine methyl ester (TMRM), here we introduce HBmito Crimson (HBmito), a novel membrane-anchored voltage sensor to probe mitochondrial near-field potential (Vnf) in intact cells by capturing fluorescence signal variations through rapid optical imaging. Vnf encompasses surface potential and the innermost portion of Δψm. During mitochondrial action potential, HBmito-reported Vnf-related signals show up to a 1.9-fold increase, concurrent with Δψm changes reported by TMRM. At an imaging rate of 1000 frames per second, transient miniature Vnf-related flickers were further revealed. And these events were three orders of magnitude smaller than action potentials and likely reflected the activity of individual mitochondrial ionic channels. These results provide the first optical imaging evidence of fundamental electrophysiological events in intact mitochondria, and highlight a novel approach for studying excitable membranes.

Recent years have witnessed great advances in applying deep learning to improve fluorescence microscopy imaging. However, enhancing the fidelity of image restoration networks and improving their robustness under fluorescence noise remain significant challenges.

Peking University, June 9, 2026: Recent years have witnessed great advances in applying deep learning to improve fluorescence microscopy imaging. However, enhancing the fidelity of image restoration n...

Existing small patch design in CNN and Transformer restoration network deviates from characteristics of fluorescence microscopy. LargePNet overcomes this context-deficiency, significantly improving st...

The crowded intracellular milieu shapes organelle architecture and dynamics, yet nanoscale heterogeneity in its physicochemical properties remains difficult to visualize with conventional fluorescence anisotropy (FA) imaging. Here, we develop fluorescence anisotropy structured illumination microscopy (FA-SIM), which employs orthogonal-polarization structured illumination with dual-angle detection to achieve ∼100-nm resolution and quantitative FA retrieval with 0.56% relative error, representing over 20-fold higher accuracy than conventional FA imaging. With low phototoxicity, FA-SIM enables dual-color, hour-long quantitative super-resolution imaging in cells. Using viscosity standards, defined nanoparticles and small-molecule drug binding assays, we validate FA-SIM as a quantitative reporter of rotational mobility and molecular interactions. In cells, FA-SIM resolves nanoscale crowding heterogeneity, correlates anisotropy landscapes with condensate dynamics, and uncovers a radial crowding gradient across the microtubule network and mitotic spindle. Long-term dual-color imaging further resolves coordinated actin–microtubule remodeling and associated microenvironmental changes. By enabling quantitative, super-resolved mapping of intracellular physical properties in living systems, FA-SIM provides a powerful platform for investigating the physical regulation of cellular organization and dynamics in health and disease. ### Competing Interest Statement Dr. Peng Xi holds the position of Chief Technology Officer (CTO) at Airy Technologies. He declares that there are no additional financial or personal interests that could be perceived as a conflict of interest in relation to the research presented in this paper. The other authors declare no competing interests. National Natural Science Foundation of China, 62025501, 31971376, 92150301, 62411540238, 62335008, 62405010 National Key R&D Program of China, 2022YFC3401100

For as long as humans have peered through lenses, the microscope has been our gateway to hidden worlds. From Robert…

Triangle-beam interference structured illumination microscopy leverages radially polarized beams to generate two-dimensional lattice illumination patterns. The technique enables a temporal resolution ...

C²SD-ISM integrates an SD and a DMD to form a dual confocal configuration, enabling deep-tissue super-resolution imaging with DPA-PR reconstruction.

A suite of bridged rhodamine dyes (BriDyes) offers excellent brightness, solubility, photostability, and tunable cell permeability along with resistance to photoblueing, making them exceptional all-pu...

A suite of bridged rhodamine dyes (BriDyes) offers excellent brightness, solubility, photostability, and tunable cell permeability along with resistance to photoblueing, making them exceptional all-pu...

Dark sectioning is a new method for removing defocused background signal in fluorescence microscopy, leveraging dark-channel priority in natural photography dehazing to substantially enhance optical s...