Abstract
Dynamic mapping of extracellular pH (pHe) at the single-cell level is critical for understanding the role of H+ in cellular and subcellular processes, with particular importance in cancer. While several pHe sensing techniques have been developed, accessing this information at the single-cell level requires improvement in sensitivity, spatial and temporal resolution. We report on a zwitterionic label-free pH nanoprobe that addresses these long-standing challenges. The probe has a sensitivity > 0.01 units, 2 ms response time, and 50 nm spatial resolution. The platform was integrated into a double-barrel nanoprobe combining pH sensing with feedback-controlled distance dependance via Scanning Ion Conductance Microscopy. This allows for the simultaneous 3D topographical imaging and pHe monitoring of living cancer cells. These classes of nanoprobes were used for real-time high spatiotemporal resolution pHe mapping at the subcellular level and revealed tumour heterogeneity of the peri-cellular environments of melanoma and breast cancer cells.
Highlights
Dynamic mapping of extracellular pH at the single-cell level is critical for understanding the role of H+ in cellular and subcellular processes, with particular importance in cancer
An example that we propose to take advantage of is in the dryingmediated self-assembly of a poly-l-lysine/ Glucose oxidase (PLL/ GOx) hydrogel, which can be crosslinked by using glutaraldehyde vapour[17]
Such nanomembranes allow the ion current to flow through the membrane matrix, which can be used for feedback control when coupled to live-cell imaging methods such as Scanning Ion Conductance Microscopy (SICM)
Summary
Dynamic mapping of extracellular pH (pHe) at the single-cell level is critical for understanding the role of H+ in cellular and subcellular processes, with particular importance in cancer. The positively charged quaternary amines of PLL and negatively charged carboxylic acid residues of GOx facilitates the self-assembly of zwitterion-like membranes, ensuring heightened H+ sensitivity Such nanomembranes allow the ion current to flow through the membrane matrix, which can be used for feedback control when coupled to live-cell imaging methods such as Scanning Ion Conductance Microscopy (SICM). We report on the development of a label-free pH-sensitive nanoprobe consisting of a self-assembled zwitterion-like nanomembrane at the tip of a nanopipette This platform allows for SICM feedback-controlled precise positioning of the nanoprobe to the cell surface to monitor the local pHe with high spatiotemporal resolution and high sensitivity. We show that double-barrel SICM-pH nanoprobes can be fabricated and used to combine the advantages of high-resolution SICM feedback-controlled scanning with high-sensitivity pH-sensing, enabling the acquisition of simultaneous topography-pHe 3D mapping of single living cells in real-time, Fig. 1a
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