Development of a fibre optic micro-optrode for intracellular pH measurements

The authors describe the production and experimental evaluation of a miniaturised fibre optic micro-optrode which has been designed for intracellular measurements. The fabrication process initially involves the controlled tapering of a single-mode optical fibre to yield micron- and submicron-diameter tips. By selectively coating the fibre tips with a thin metal film, it is possible to form a light-transmissive optical aperture at the tip. Such tapered and coated fibres are similar to the probes used in near-field optical microscopy. The probes here are converted to micron and submicron pH sensors by immobilising a fluorescent pH sensitive reagent within a glass film which is deposited on the surface of the fibre tip using a sol-gel process. When appropriate pump radiation is guided to the tip of the sensor, the fluorescence yield of the pi-I-sensitive reagent is indicative of the pH of the environment of the sensor. The authors have successfully used these sensors to monitor the pH of buffer solutions inside micron-size holes in a polycarbonate membrane and to probe the intracellular environment of mouse embryonic fibroblast cells. It was found that the sensors were reversible in the pH range 3-10.