Highly selective and stable microdisc biosensors for l-glutamate monitoring

Abstract

a b s t r a c tGlutamate mediates most of the excitatory synaptic transmission in the brain, and its abnormal regulation is considered a key factor underlying the appearance and progression of many neurodegenerative and psychiatric diseases. In this work, a microdisc-based amperometric biosensor for glutamate detection with highly enhanced selectivity and good stability is proposed. The biosensor utilizes the enzyme glutamate oxidase which was dip-coated onto 125 m diameter platinum discs. To improve selectivity, phosphatidylethanolamine was pre-coated prior to enzyme deposition, and electropolymerization of o-phenylenediamine was performed to entrap the enzyme within a polymer matrix. A variety of coating configurations were tested in order to optimize biosensor performance. For stability measurements, biosensors were biased continuously and calibration curves calculated each day for a period of 5-6 days. The optimized biosensors exhibited very high sensitivity (71 ± 1 mA M −1 cm −2 ), low detection limit of ∼2.5 M glutamate, selectivity (over 87% against ascorbic acid), very good temporal stability during continuous use, and a response time of <5 s. These biosensors are therefore good candidates for further development as devices for continuous monitoring during traumatic brain injury or neurosurgery.

Characterization of Biosensors Based on Recombinant Glutamate Oxidase: Comparison of Crosslinking Agents in Terms of Enzyme Loading and Efficiency Parameters

“…This is particularly important when the enzyme substrate is anionic, as in the case of Glu detection at pH values close to neutrality [47]. In the absence of PEI, a high surface loading of wild-type GluOx has been shown to result in a high K M value, i.e., a low affinity of Glu for the enzyme, which was reversed by surface PEI [26,48]. The balance of PEI and recombinant GluOx loading was therefore investigated here to provide the best analytical parameters (especially sensitivity and stability) for long-term Glu monitoring.…”

“…Many previously reported descriptions of PEI-containing Glu biosensors were based on wild-type GluOx [26,49,77,78]. Since recombinant enzyme is now more readily available [53], a comparison of biosensor parameters for native vs. recombinant GluOx was undertaken.…”

“…Moreover, long-term in vivo electrochemistry (LIVE) has been demonstrated in the case of sensors, such as carbon-fiber or carbon-paste electrodes for monitoring neurotransmitters [23] and their metabolites [24] over periods of months. However, the inherent complexity of biosensors, incorporating a range of components (including enzymes, permselective membranes and enzyme stabilizers), aggravates the challenge of their long-term stability in biological tissues and other complex media [24,25,26]. In this work, a biosensor was designed for the detection of Glu, with an emphasis on sensitivity and stability, based on both wild-type and recombinant l -glutamate oxidase (GluOx) as the sensing element.…”

“…Here, we build on previous studies of PoPD-modified, wild-type GluOx-based biosensors designed for LIVE applications which have demonstrated, in addition to outstanding permselectivity, the following features: electrostatic stabilization [47] of GluOx with polycationic polyethyleneimine, PEI [26,48,49]; low limits of detection (<1 µM) [50,51]; fast response times (~1 s) [51]; and their negligible oxygen dependence (Equation (2)) for concentrations of Glu and O 2 relevant to neurochemical monitoring [48,50]. However, their long-term stability was limited (days) [52].…”

Recent Developments in Electrochemical Sensors for the Detection of Neurotransmitters for Applications in Biomedicine

“…Platinum electrodes provide the most sensitive detection for H 2 O 2 due to the electocatalytic effect of Pt on H 2 O 2 oxidation (Chen 2012). To prevent interferences from other electroactive compounds (specifically ascorbic acid (∼200 µM), uric acid, dopamine, and serotonin) that are present in the physiological media, electrodes were casted by perm-selective materials such as Nafion (Pan 1996, Govindarajan 2013), o -phenylenediamine (Lowry 1998, R. Ryan 1997, Özel 2014), over-oxidized polypyrrole (Walker 2007, Hamdi 2006) and other non-traditional membranes (Mizutani 1998, McMahon 2006).…”