Glutaraldehyde Cross-Linked Glutamate Oxidase Coated Microelectrode Arrays: Selectivity and Resting Levels of Glutamate in the CNS

Glutaraldehyde is widely used as a cross-linking agent for enzyme immobilization onto microelectrodes. Recent studies and prior reports indicate changes in enzyme activity and selectivity with certain glutaraldehyde cross-linking procedures that may jeopardize the performance of microelectrode recor...

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Published in:ACS chemical neuroscience 2013-05, Vol.4 (5), p.721-728
Main Authors: Burmeister, Jason J, Davis, Verda A, Quintero, Jorge E, Pomerleau, Francois, Huettl, Peter, Gerhardt, Greg A
Format: Article
Language:English
Subjects:
Amino Acid Oxidoreductases - drug effects
Amino Acid Oxidoreductases - physiology
Biosensing Techniques
Cross-Linking Reagents - pharmacology
Enzymes, Immobilized - drug effects
Enzymes, Immobilized - physiology
Glutamic Acid - analysis
Glutaral - pharmacology
Letter
Microelectrodes
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Summary:Glutaraldehyde is widely used as a cross-linking agent for enzyme immobilization onto microelectrodes. Recent studies and prior reports indicate changes in enzyme activity and selectivity with certain glutaraldehyde cross-linking procedures that may jeopardize the performance of microelectrode recordings and lead to falsely elevated responses in biological systems. In this study, the sensitivity of glutaraldehyde cross-linked glutamate oxidase-based microelectrode arrays to 22 amino acids was tested and compared to glutamate. As expected, responses to electroactive amino acids (Cys, Tyr, Trp) were detected at both nonenzyme-coated and enzyme-coated microelectrodes sites, while the remaining amino acids yielded no detectable responses. Electroactive amino acids were effectively blocked with a m-phenylene diamine (mPD) layer and, subsequently, no responses were detected. Preliminary results on the use of poly­(ethylene glycol) diglycidyl ether (PEGDE) as a potentially more reliable cross-linking agent for the immobilization of glutamate oxidase onto ceramic-based microelectrode arrays are reported and show no significant advantages over glutaraldehyde as we observe comparable selectivities and responses. These results support that glutaraldehyde-cross-linked glutamate oxidase retains sufficient enzyme specificity for accurate in vivo brain measures of tonic and phasic glutamate levels when immobilized using specific “wet” coating procedures.
ISSN:1948-7193
1948-7193
DOI:10.1021/cn4000555