On-Chip Amperometric Measurement of Quantal Catecholamine Release Using Transparent Indium Tin Oxide Electrodes

Abstract

Carbon-fiber amperometry has been extensively used to monitor the time course of catecholamine release from cells as individual secretory granules discharge their contents during the process of quantal exocytosis, but microfabricated devices offer the promise of higher throughput. Here we report development of a microchip device that uses transparent indium tin oxide (ITO) electrodes to measure quantal exocytosis from cells in microfluidic channels. ITO films on a glass substrate were patterned as 20-mum-wide stripes using photolithography and wet etching and then coated with polylysine to facilitate cell adherence. Microfluidic channels (100 mum wide by 100 mum deep) were formed by molding poly(dimethylsiloxane) (PDMS) on photoresist and then reversibly sealing the PDMS slab to the ITO-glass substrate. Bovine adrenal chromaffin cells were loaded into the microfluidic channel and adhered to the ITO electrodes. Cells were stimulated to secrete by perfusing a depolarizing "high-K" solution while monitoring oxidation of catecholamines on the ITO electrode beneath the cell using amperometry. Amperometric spikes with charges ranging from 0.1 to 1.5 pC were recorded with a signal-to-noise ratio comparable to that of carbon-fiber electrodes. Further development of this approach will enable high-throughput measurement of quantal catecholamine release simultaneously with optical cell measurements such as fluorescence.

Electrochemical Probes for Detection and Analysis of Exocytosis and Vesicles

“…They found that amperometric exocytosis events were greatest where the calcium level was highest on the other side of the plasma membrane. [43] In the quest to improve spatial resolution recording, thin platinum [17] and indium tin oxide [19] electrodes have been patterned on surfaces for electrochemical detection of exocytosis from single cells. These methods also allow simultaneous microscopy imaging.…”

Release monitoring of single cells on a microfluidic device coupled with fluorescence microscopy and electrochemistry

“…[1][2][3] An alternative way to study the process of exocytotic release is by electrical measurements that mainly include electrophysiological ͑i.e., membrane capacitance measurements͒ 4,5 and electrochemical detection ͑e.g., constant-potential amperometry, high-speed chronoamperometry, fastscan cyclic voltammetry͒. [6][7][8][9][10] Another possible approach to study exocytosis is through optical methods. [11][12][13][14][15] Among them, amperometry using microelectrodes and fluorescence microscopy especially in an evanescent wave field are considered to be two of the most powerful techniques.…”

On‐Chip Determination of Dopamine Exocytosis Using Mercaptopropionic Acid Modified Microelectrodes

“…A few examples of microchip systems for measurement of quantal exocytosis can be found in the literature. Measurement of adrenaline and noradrenalin exocytosis from Chromaffin cells was demonstrated by Chen et al in a gold plated picoliter well [11], by Sun et al in a microfluidic system containing a transparent ITO electrode [12] while Dias et al used a Pt array on glass [13]. DA exocytosis from PC12 cells has been measured in a microfluidic system containing a Pt-electrode [14] whereas Huang et al used a microfluidic system to position a cell prior to measurement using a carbon fiber microelectrode [5].…”