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We report on the effect of depositing the ionophore valinomycin onto the polymethylmethacrylate (PMMA) gate insulator of an ion-sensitive organic field-effect transistor (ISOFET) based on poly(3-hexylthiophene). The ionophore was deposited onto the PMMA using the Langmuir-Blodgett (LB) technique; thin films based on pure valinomycin and those in which valinomycin was mixed with arachidic acid were investigated. The pH sensitivity of the reference ISOFET could be improved significantly when the devices were coated with an LB film of arachidic acid. However, the response to was low. By adding a small amount (5% w/w) of the ionophore valinomycin to the fatty acid LB film, an improved response to potassium ions was achieved, but no selectivity over sodium. It was necessary to use an LB membrane of pure valinomycin in order to realise an ISOFET with some selectivity. We suggest that the presence of the ionophore in the fatty acid matrix disrupts the packing of the hydrocarbon chains in the mixed LB film and that the monovalent ion response originates from interactions with the carboxylic acid groups in the fatty acid. In contrast, for the case of the pure valinomycin coating, it is thought that response is controlled by complex formation with the ionophore.