Near-Field Imaging of Cell Membranes in Liquid Enabled by Active Scanning Probe Mechanical Resonance Control

Despite the power of far-field super-resolution microscopies for three-dimensional imaging of biomolecular structures and processes, its application is challenged in dense and crowded samples and for certain surface and membrane studies. Although near-field imaging with its ability to provide intrinsic subdiffraction limited spatial resolution at any optical modality, its application to biological systems has remained limited because of the difficulties of routine operation in liquid environments. Here we demonstrate stable and sensitive near-field scanning optical microscopy (NSOM) in a liquid based on a new mechanical resonance control and an optimization of the tip length, achieving a high quality factor (>2800) force sensing of the near-field probe. Through near-field imaging of the spatial distribution of epidermal growth factor receptors (EGFRs) on the membrane of A431 cancer cells as an example, we reveal nanoscale correlations between surface EGFR and intracellular organelle structures with ∼50 nm spatial resolution. The method provides a new avenue for surface imaging in viscous liquid media to complement super-resolution microscopy for studies of biological membranes, nanostructures, and interfaces.