title: Image processing for non-ratiometric measurement of membrane voltage using fluorescent reporters and pulsed laser illumination.
abstract: The measurement of transmembrane voltages induced by pulsed electric field exposure can be achieved by using fluorescent dyes like ANNINE-6. Such approach requires a quantitative determination of the fluorescence intensity along the cell's membrane by image processing. When high temporal resolution is required, the illumination source is frequently a dye-laser which causes high fluctuations in the intensity of illumination which in turn affects the fluorescence intensity and thus the quality of the results. We propose an image processing technique that allows to overcome the fluctuations and to produce quantitative data. It uses the optical background noise as a correcting factor. Standard deviation in the fluctuations is thus efficiently reduced by at least a factor of 2.5. Additionally we draw attention to the fact that the parasitic component of the laser radiation (ASE) can also suppress fluctuations although it deteriorates wavelength precision.
authors.affiliation: Institute for Pulsed Power and Microwave Technology, Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, Eggenstein-Leopoldshafen 76344, Germany. Electronic address: aude.silve@kit.edu.
authors.forename: Aude
authors.initials: A
authors.lastname: Silve
authors.affiliation: Institute for Pulsed Power and Microwave Technology, Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, Eggenstein-Leopoldshafen 76344, Germany.
authors.forename: Sarah
authors.initials: S
authors.lastname: Rocke
authors.affiliation: Institute for Pulsed Power and Microwave Technology, Karlsruhe Institute of Technology, Hermann-von-Helmholtz-Platz 1, Eggenstein-Leopoldshafen 76344, Germany. Electronic address: wolfgang.frey@kit.edu.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Frey
mesh_terms: D000476:Alkanesulfonates; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism; D002911:Chrysenes; D003412:Cricetulus; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000032:analysis; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D007834:Lasers; D059629:Signal-To-Noise Ratio
chemical_list: C000606998:ANNINE-6; D000476:Alkanesulfonates; D002911:Chrysenes; D005456:Fluorescent Dyes
keywords: ANNINE-6; Image processing; Intensity fluctuations; Transmembrane potential; Transmembrane voltage