Development and Evaluation of a Microwire Biosensor for the Detection of Fumarate

Enzymatic microwire biosensors offer potential advantages in real-time, in-situ biological analyses due to their high sensitivity, selectivity, and miniaturisation capabilities. In this study, we present the design, fabrication, and evaluation of a fumarate-specific microwire biosensor intended for potential in-situ measurements in cerebrospinal fluid. The sensor employs a two-electrode self-referential setup and a cascade enzymatic reaction involving fumarase and malate dehydrogenase immobilised on a gold-coated tungsten microwire, with Nafion used for enzyme immobilisation and as a protective membrane. This innovative design allows for selective detection of fumarate, an important metabolite in the Krebs cycle and implicated in neurodegenerative diseases, including multiple sclerosis. Our fabricated biosensor demonstrates consistent electrochemical responses across a range of fumarate concentrations, with minimal interference from similar compounds such as succinate and malate. These findings validate the sensor's sensitivity and selectivity, highlighting its potential applicability for in-situ measurements in complex biological environments. Future work will focus on enhancing sensor longevity for long-term in-vivo applications, evaluating performance in ex-vivo mouse cerebrospinal fluid samples, and ultimately validating its functionality through in-situ testing in a live mouse model. The successful implementation of such a sensor could provide invaluable insights into metabolite dynamics in the brain, contributing to our understanding of physiological and pathological processes in neurodegenerative diseases.