Multiparametric Correlative Topographical and Volumetric Fluorescence Microscopy

Abstract

Live-cell imaging of cell surface topography and intracellular architecture is essential for understanding cellular function. However, conventional approaches often involve trade-offs between resolution, invasiveness, and volumetric coverage. Here, we present an integrated Scanning Ion Conductance Microscope and single-objective Oblique Plane Microscope (SICM-OPM) system that enables simultaneous non-contact topographical imaging and volumetric fluorescence imaging within the same live cell. Beyond correlative live imaging, the platform supports nanomechanical mapping with tens-of-nanometres resolution, fluorescence-guided localised molecular delivery via the SICM, and benefits from reduced photobleaching due to light-sheet excitation. We demonstrate this platforms capabilities by visualising imipramine-induced T-tubule remodelling in live cardiomyocytes, revealing subsurface detubulation while surface morphology remains preserved. Additionally, we show precision delivery of fluorescent cargos--including dextrans and -synuclein--into diatom and mammalian cells, alongside localised stiffness mapping to evaluate mechanical responses. We believe this technique opens new avenues for correlative structural, functional, and biophysical studies in live cells, with broad relevance to cell biology, neurodegeneration, and mechanobiology.