In order to detect L-glutamate, we immobilized L-glutamate oxidase (GLOD) on the probe-type carbon nanotube (CNT) network junctions by a non-covalent functionalized method to preserve their electronic characteristics. After immobilizing GLOD on the CNT network, the excess reactive groups of linker molecule remaining on its surface were deactivated and blocked by ethanolamine. The electrical property of the GLOD-immobilized CNT network transistor was characterized as source-drain current that depended on liquid gate voltage. The real time electronic response of the GLOD-coated CNT network junctions was conducted with a glutamate standard solution in vitro and the 11 vessel occlusion (11VO) rat model in vivo. The ultrahigh sensitivity, selectivity, and fast response time of GLOD-immobilized CNT-FET (field-effect transistor) could provide great potential for the real time electronic detection of extracellular glutamate levels in the brain.