BPS2026 – Time-resolved high-pressure freezing with ligand stimulation

Imaging cellular dynamics requires millisecond temporal resolution and nanometer spatial resolution—a blind spot of most existing microscopes. To address this need, we developed a high-pressure freezing instrument, high pressure freezing with buffer exchange (HPF-X). HPF-X enables rapid chemical stimulation of cells and vitrification within ∼500 ms following stimulation. This method arrests early time points of cellular processes and allows for closer inspection of transient events using high-resolution imaging methods such as super-resolution fluorescence microscopy and cryo-electron microscopy. By vitrifying replicate samples at sequential time points, HPF-X permits reconstructing a montage of rapid ultrastructural and molecular motions. As a proof of principle, we applied HPF-X to study the growth dynamics of biomolecular condensates induced by hyperosmotic stress. HPF-X has the potential to find broad utility for investigating dynamic processes in single cells and tissues.