7 October 2005 TIR fluorescence reader for selective detection of cell membranes
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Abstract
A novel setup for fluorescence measurements of surfaces of biological samples, in particular cell membranes, is described. The method is based on multiple total internal reflections (TIR) of a laser beam at the surface of a multi-well plate, such that 96 individual samples are excited simultaneously. Main prerequisites are an appropriate thickness and high transmission of the glass bottom, a non-cytotoxic adhesive, and appropriate glass rods for TIR illumination. Fluorescence from the cell surface is detected simultaneously using an integrating CCD camera and appropriate optical filters. For validation of the system, cells incubated with the fluorescence marker NBD as well as transfected cells expressing a fluorescent membrane protein are used. In addition, intracellular translocation of a fluorescent protein kinase c fusion protein upon stimulation is examined. The method appears well suitable for high throughput screening (HTS), since neither washing of the samples nor any readjustment of the equipment after changing of individual plates are necessary.
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Thomas Bruns, Wolfgang S. L. Strauss, Reinhard Sailer, Michael Wagner, Herbert Schneckenburger, "TIR fluorescence reader for selective detection of cell membranes", Proc. SPIE 5859, Photon Migration and Diffuse-Light Imaging II, 585908 (7 October 2005); doi: 10.1117/12.632828; https://doi.org/10.1117/12.632828
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