Fig. 1.

Nanoprobe pH sensor fabrication and characterisation. a Conceptual image demonstrating the dynamic mapping of extracellular pH in three dimensions with high spatial resolution. b The nanoprobe pH sensors were made by immobilising a mixture of glucose oxidase and poly-l-lysine and made via drying-mediated self-assembly at the tip of a pulled glass nanopipette by crosslinking with glutaraldehyde. c A scanning electron microscopy image (scale bar 500 nm) showing the tip of the pH probe consisting of a thin membrane. The pipette tip has been focused ion beam milled using a slice and view technique. d Proposed working principle of the sensor, by which the nanomembrane shows preferential permeability for anions at low-pH and cations at high-pH. e Current-voltage characterisation of the sensor at varying pH, and f current vs pH at 0.6 V showing good linear response in the pH range of 4–9 (R² = 0.96, p < 0.001, Pearson’s correlation). g A nanopipette was used as a highly localised H⁺ source for testing the pH mapping capability of the nanoprobe sensor (left top inset). 2D top view showing the pH distribution profile (right bottom inset) as obtained using SICM mapping. Real-time pH measurements that allow assessing the probe response time and sensitivity. The probe is moved to the H⁺ source in the z-direction using a fast piezo-stage. h A magnified plot of the dotted-box shown in g, demonstrating the sensitivity and resolution of the nanoprobe pH sensor.