Electrochemical Bioimaging Platform Using Closed Bipolar Electrode Array

Abstract

Introduction Bio-imaging technologies have greatly contributed to the fields of life science and therapeutic researches by providing information about functions of cells and living tissues. Electrochemical imaging systems with micro/nano electrode can provide local chemical information without disturbing the bio-system by label molecules, which contributes to the understanding of intercellular communication of multicellular organism. Conventional electrochemical imaging systems is classified into two types: One is scanning electrochemical microscopy (SECM) and the other is electrode array chip. SECM can provide spatially high-resolution images by scanning an ultramicroelectrode probe. However, time resolution of SECM is not so high due to the time required to scan the probe. Electrode array method has an advantage in time resolution which allows real-time observation. However, conventional electrode arrays require the presence of connecting lines between the voltage supplier and individual electrodes, which severely limit the spatial resolution.In this study, to realize the real-time bioimaging with single-cell level spacial resolution, we have employed a closed bipolar electrode (cBPE) array with electrochemiluminescence (ECL) detecting system as a new platform for electrochemical imaging [1]. Our cBPE array electrochemical imaging system is composed of a bipolar electrode (BPE) array that connects two separated electrochemical cells, with a pair of driving electrodes in each cell. By applying an adequate potential between the two driving electrodes, redox reactions at the BPEs in the sample cell can be detected by the detection cell with the ECL. Since the BPE does not require any connecting lines to work, arrays with a large number of electrodes can be easily integrated into the platform with high density. Experimental As a principal verification device, we fabricated a cBPE array by bundling Au wires (200 µm in diameter), each of them is coated with epoxy resin (Fig. A). The cBPE array was put between the sample cell and detection cell. An Ag/AgCl reference electrode was lowered into the sample cell, and the Pt driving electrodes in the sample cell and detection cell were connected to the working electrode and counter electrode terminals of the potentiostat, respectively. In the detection cell, we put 10 mM [Ru(bpy)3]2Cl2 with 100 mM tripropylamine (TPA) in a 0.1 M KCl solution as a luminophore for ECL. To demonstrate the chemical imaging, 500 µL of a 50 mM K3[Fe(CN)6] containing 0.1 M KCl solution was injected into a 0.1 M KCl solution in the sample cell, while applying a −1.1 V to the driving electrode and observing ECL from the bottom side of detecting cell by CCD camera (Fig. B). We also fabricated higher density cBPE array for higher special resolution imaging. Pores of track-etched membrane (8 µm in diameter) was filled with gold by electroless deposition using 50 mM chlorauric acid and 50 mM NaBH4. Result and discussion As shown in Fig. C, we successfully obtained a real-time image of the flow of the injected K3[Fe(CN)6] as ECL signal using the principal verification device. When [Fe(CN)6]3− was reached to one end of a BPE in the sample cell, the [Fe(CN)6]3− was reduced to [Fe(CN)6]4− and Ru/TPA were oxidized at another end of BPE in the detection cell resulting in the production of ECL signal. We also successfully obtained a flow image of [Fe(CN)6]3− using the higher density cBPE array with an average pixel distance of 35 µm. The results indicate that the proposed cBPE array system can be widely applied in bio/chemical imaging. The future aim is to improve the cBPE array by modulating its diameter and density of the electrodes for high-resolution imaging, which may result in the development of a novel technology for its application in chemical imaging, including single-cell bioimaging.