Regensburg 2010 – scientific programme

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BP: Fachverband Biologische Physik

BP 2: New Technologies

BP 2.2: Talk

Monday, March 22, 2010, 10:45–11:00, H43

Near-fields in Fluorescence Microscopy - Absolute Determination of z-positions in the Nanometer Range — •Michael Berndt¹, Mike Lorenz¹, Jörg Enderlein², and Stefan Diez¹ — ¹Max-Planck-Institute of Molecular Cell Biology and Genetics, Dresden, Germany — ²III. Institute of Physics, Georg-August-University, Göttingen, Germany

Typical processes in molecular biology take place on a nanometer range and are too small to be resolved by conventional light microscopy. To though reveal information from such systems, techniques are required which access information on the nanometer scale. Here, we present a novel near-field based method to precisely determine absolute heights on the length scale of 0-100 nm above surfaces. We make use of distance dependent quenching processes fluorophores undergo in the proximity to conductive interfaces and measure the influence on the fluorescence lifetime by wide-field fluorescence lifetime imaging microscopy [1]. We use theoretical computations based on the CPS theory to scale the determined lifetimes to absolute heights [2]. We apply our method to fluorescently labeled microtubules elevated to different heights by several spacer molecules. Hence we demonstrate a technique to measure molecule geometries with nanometer precision.

References: [1] M. Lorenz, RNA 15, 97-103 (2009), [2] R.R. Chance, A. Prock, R. Silbey, Advances in Chemical Physics XXXVII, 1-65 (1978)