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

Abstract

A method for monitoring the biological exocytotic phenomena on a microfluidic system was proposed. A microfluidic device coupled with functionalities of fluorescence imaging and amperometric detection has been developed to enable the real-time monitoring of the exocytotic events. Exocytotic release of single SH-SY5Y neuroblastoma cells was studied. By staining the cells located on integrated microelectrodes with naphthalene-2,3-dicarboxaldehyde, punctuate fluorescence consistent with localization of neurotransmitters stored in vesicles was obtained. The stimulated exocytotic release was successfully observed at the surface of SH-SY5Y cells without refitting the commercial inverted fluorescence microscope. Spatially and temporally resolved exocytotic events from single cells on a microfluidic device were visualized in real time using fluorescence microscopy and were amperometrically recorded by the electrochemical system simultaneously. This coupled technique is simple and is hoped to provide new insights into the mechanisms responsible for the kinetics of exocytosis.

Fabrication of two-layer poly(dimethyl siloxane) devices for hydrodynamic cell trapping and exocytosis measurement with integrated indium tin oxide microelectrodes arrays

“…12, 13 A microfluidic device coupled with functionalities of fluorescence imaging and amperometric detection has been developed to enable the real-time monitoring of the exocytotic events of single SHSY5Y neuroblastoma cells. 14 Sigworth’s group at Yale fabricated poly(dimethyl siloxane) (PDMS) substrates to replace the pipette and successfully recorded single ion channel current from Xenopus oocytes. 15 Fertig and colleagues reported a planar patch-clamp glass chip to record electrical current measure from different cell types in a whole-cell mode.…”

Orthogonal Cell‐Based Biosensing: Fluorescent, Electrochemical, and Colorimetric Detection with Silica‐Immobilized Cellular Communities Integrated with an ITO–Glass/Plastic Laminate Cartridge

“…In these pioneering works exocytosis from chromaffin cells was monitored under conditions of continuous perfusion culture. Although ideal for fundamental molecular biology studies performed over several hours these, and other similar devices,13–15 were not designed for long‐term cell‐based environmental biosensing. In another recent study, the utility of coupling electrochemical and fluorescent techniques for cell‐based biosensing of pesticides was reported 16.…”

Emission Spectroelectrochemistry: Cell Design and Setup

“…Indeed, Amatore et al [64] have reported the real-time observation of fluorescence and amperometric measurements of exocytosis events at single biologic cell on transparent indium tin oxide (ITO) microelectrodes(Figure 12). Similar investigations were done using microfluidic configuration[65].…”