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Amperometric hydrogel-coated glutamate microsensors form a promising concept to detect glutamate levels directly in brain tissue. These microsensors are constructed by coating a carbon fiber electrode (CFE) (10 microm diameter; 300-500 microm long) with a five-component redox-hydrogel, in which L-glutamate oxidase, horseradish peroxidase, and ascorbate oxidase are wired via poly(ethylene glycol) diglycidyl ether to an osmium-containing redox polymer. Coating with a thin Nafion film completes the construction. Prior to use in vivo, a reliable and reproducible construction of microsensors with a high performance is required. For an optimal microsensor performance, the balance between the five individual hydrogel components is critical. However, due to their small size, hydrogel application to CFE's need to be performed by dip-coating. Dip-coating is a difficult procedure to control and does not allow individual application of hydrogel constituents. To improve the microsensor construction and to better control the dip-coating procedure, we have recently developed an automated device. Throughout this study, automatic dip-coating was performed with premixed solutions, in which the amount of a single component was varied. This allowed us to optimize the hydrogel composition, which resulted in a significant improvement of the microsensor properties in terms of sensitivity, current density, linearity, detection limit, and interference by ascorbic acid.

Glutamate microsensors form a promising analytical tool for monitoring neuronally derived glutamate directly in the brain. However, when a microsensor is implanted in brain tissue, many factors can diminish its performance. Consequently, a thorough characterization and evaluation of a microsensor is required concerning all factors that may possibly be encountered in vivo. The present report deals with the validation of a hydrogel-coated glutamate microsensor. This microsensor is constructed by coating a carbon fiber electrode (10-microm diameter; 300-500 microm long) with a five-component redox hydrogel, in which L-glutamate oxidase, horseradish peroxidase, and ascorbate oxidase are wired via poly(ethylene glycol) diglycidyl ether to an osmium-containing redox polymer. A thin Nafion coating completes the construction. Although this microsensor was previously used in vivo, information concerning its validation is limited. In the present study, attention was given to its selectivity, specificity, calibration, oxygen dependency, biofouling, operating potential dependency, and linear range. In addition, successful microsensor experiments in microdialysate, in vitro (in organotypic hippocampal slice cultures), and in vivo (in anesthesized rats) are shown.

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