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A two-photon electrochemical and fluorescence microscope for imaging of cell-surface interactions (FLUOLAPS)
Start date: 01 Sep 2014, End date: 31 Aug 2016 PROJECT  FINISHED 

Traditional microscopy provides information about structure, but is blind to functional dielectric and electric properties of a system. In contrast, microelectrode arrays, which can provide high sensitivity electrophysiological recordings, are difficult to combine with imaging. A two-photon electrochemical and fluorescence microscope capable of producing high-resolution, two-dimensional electrochemical images of parameters such as extracellular potentials, surface charges and impedance and two-photon fluorescence images of the cell-attachment area simultaneously will be developed and used to investigate cell-surface interactions and cell signalling. The instrument will incorporate the capabilities of the impedance imaging technique, Scanning Photo-induced Impedance Microscopy (SPIM), Light-Addressable Potentiometric Sensors (LAPS) and two–photon fluorescence microscopy. The proposed technology has a unique twist in that the semiconductor substrate used as the LAPS/SPIM substrate also serves as the fluorescence detector providing a simple, elegant solution to a complex measurement problem. As the laser beams used for the excitation of LAPS/SPIM and fluorescence signals are focused through the same microscope objective, we will, for the first time, be able to access the same micro-environment with two photon fluorescence microscopy and electrical imaging simultaneously, enabling us to monitor dynamic changes in cell morphology and electrical properties in real time. The strengths of the technique lie in the fact, that cell-surface interactions can be investigated on any material that can be deposited onto a semiconductor substrate, that any point on the substrate can be addressed with a focused laser beam, i.e. the resolution is not limited by the miniaturisation of an electrode or transistor array, and LAPS and SPIM measurements can be performed with high sensitivity due to the use of an organic monolayer as the insulator.
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