A novel light source for SICM–SNOM of living cells

We have developed a novel light source for use in a scanning near‐field optical microscope (SNOM or NSOM) based on a nanopipette whose distance from the sample surface is controlled using scanning ion conductance microscopy. The light source is based on the general principle of the chemical reaction between a fluorophore in the pipette and ligand in the bath, to produce a highly fluorescent complex that is continually renewed at the pipette tip. In these experiments we used fluo‐3 and calcium, respectively. This complex is then excited with an Ar⁺ laser, focused on the pipette tip, to produce the light source. This method overcomes the transmission problem of more traditional SNOM probes and has been used to acquire simultaneous high‐resolution topographic and optical images of biological samples in physiological buffer. A resolution of ～220 nm topographic and ～190 nm optical was determined through imaging fixed sea‐urchin sperm flagella. Live A6 cells were also imaged, demonstrating the potential of this system for SNOM imaging of living cells.