Covalent Immobilization of Hydrogels on Carbon Fiber Microelectrodes for the Determination of Dopamine in the Living Mouse Brain

Real-time and in situ monitoring of dynamic changes in brain neurotransmitters is of significant importance for understanding their pathophysiological regulatory mechanisms. This study employed an in situ hydrogel covalent immobilization strategy mediated by oxidized poly-tannic acid (pTAox) to modify the carbon fiber electrode (CFE), in which the click chemical reaction between pTAox and five thiol molecules of varying chain lengths enables the oriented fixation of a seeding layer, followed by the construction of an in situ hydrogel coating via hydrogen bonding interactions with negatively charged tannic acid. It was found that the long-chain SH-PEG44-OH exhibited outstanding anti-fouling stability and interference resistance while maintaining high sensitivity, and the short-chain β-mercaptoethylamine (MEA), which displayed superior selectivity for positively charged dopamine (DA), was selected as the sealing agent. The TmPEG44-pTA/CFE microelectrode constructed based on an optimal combination exhibited outstanding selectivity, sensitivity, anti-fouling stability, and biocompatibility, as well as excellent linearity across DA concentrations ranging from 0.02 to 0.5 μM and 0.5 to 20 μM, with a detection limit of 18.9 nM (S/N = 3). Furthermore, TmPEG44-pTA/CFE facilitated real-time dynamic monitoring of DA release in the striatum of a living mouse. The present study proposes a systematic screening paradigm for the rational design of functionalized electrode interfaces, thus extending the application of the oriented covalent immobilization in situ hydrogel coating strategy to in vivo neurochemical analysis.