Analytica Chimica Acta 525 (2004) 75–82 Amperometric biosensor-based flow-through microdetector for microdialysis applications Szilveszter G´ asp´ ar a,∗ , Xiaowen Wang b , Hiroaki Suzuki b , Elisabeth Cs ¨ oregi a a Department of Biotechnology, Lund University, PO Box 124, 22100 Lund, Sweden b Institute of Materials Science, University of Tsukuba, 1-1-1 Ten-nodai, Tsukuba, Ibaraki 305-8573, Japan Received 15 April 2004; received in revised form 7 July 2004; accepted 7 July 2004 Available online 11 September 2004 Abstract A new flow-through electrochemical microcell was fabricated, in order to be used as an on-line detector in microdialysis-based investigations. The microcell is obtained by sandwich-like assembling the electrode system patterned on a glass chip and a silicon rubber gasket bearing the fluidic elements (inlet, outlet, flow channel, and containers for reference and counter electrode). Four Pt microelectrodes (100 m × 100 m squares), a Pt counter electrode, and a Ag reference electrode were patterned on a 9 mm × 14 mm glass chip by thin film technology. The silicon rubber cover was fabricated using a polydimethylsiloxane based precursor and a plastic mold, and it avoids significant dilution of the sample before getting in contact with the sensing electrodes. The microdetector can be directly connected to the outlet of the microdialysis probe reducing significantly the delay between sampling and detection. The microelectrodes were modified with enzyme-based chemistries in order to detect glucose, glutamate, and choline by electrooxidation of the hydrogen peroxide produced in the reaction of the analytes with their corresponding oxidases. Problems concerning appropriate linear range, interference elimination, and cross-talk elimination were addressed. © 2004 Elsevier B.V. All rights reserved. Keywords: Biosensors; Electrochemical microcell; Microdetector; Microdialysis 1. Introduction Since its first in vivo application 30 years ago [1], micro- dialysis has become one of the most used in vivo sampling methods. Its main application is still in monitoring biochem- ical changes in animal brains but it was already successfully used to sample from blood [2], adipose tissue [3,4], or muscle [5], and recently also from humans [6]. There are several advantages of using microdialysis-based sampling such as (i) relatively good spatial resolution when compared to other in vivo sampling methods, (ii) samples free of large biomolecules, (iii) no fluid is removed from the sur- rounding tissue, thus, causing a minimal perturbation of the studied system, and (iv) the possibility of drug administration by reverse microdialysis. ∗ Corresponding author. Tel.: +46 46 2228098; fax: +46 46 2224713. E-mail address: szilveszter.gaspar@biotek.lu.se (S. G´ asp´ ar). To take full advantage of microdialysis as a sampling method, it has to be combined with a fast, sensitive, and equally important, continuous, detection method. In the sim- plest approaches, traditional analytical methods are inter- faced with microdialysis sampling by using sample collec- tors. High performance liquid chromatography [7], radioim- munoassay [4], and nuclear magnetic resonance (NMR) [8] are only few examples of analytical methods used as off-line detectors in microdialysis experiments. In reality, often a much better time resolution is needed, than the one achieved with these off-line analytical proce- dures. Therefore, recent trend considers on-line detection with microdialysis sampling. Liquid chromatography [9,10], capillary electrophoresis [11,12], chemiluminescence detec- tors [13], enzyme fluorometric detectors [14,15], and mass spectrometry [16] were all used as on-line detection princi- ples connected to microdialysis sampling. Even though the time resolution achieved with on-line detection is much better 0003-2670/$ – see front matter © 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.aca.2004.07.041