Berlin 2012 – wissenschaftliches Programm

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BP: Fachverband Biologische Physik

BP 28: Posters: Imaging

BP 28.10: Poster

Donnerstag, 29. März 2012, 17:30–19:30, Poster A

(contribution withdrawn) Approaches for improved dual color Photoactivated Localization Microscopy — •Paolo Annibale, Mattia Greco, Marco Scarselli, and Aleksandra Radenovic — LBEN, IBI EPFL Lausanne 1015 Switzerland

Dual color PhotoActivated Localization Microscopy (PALM) presents unique challenges due to the specific photo-physical behavior of the fluorescent proteins used as a tags and to the asynchronous nature of the photoactivation and localization of the molecules belonging to each fluorophore pair. We systematically investigate the effect of molecular photoblinking and fluorescence dark times on a typical dual color PALM experiment. This allows us to propose a method to identify potential photoblinking-originated artifacts by screening for concomitant spatial and temporal clusters, a prerequisite for the correct localization and co-localization of oligomeric sub-diffraction limit cellular structures [1]. Using a setup axially stabilized to better than 5 nm we compare the performance of three different dual color fluorescent protein pairs using as positive controls fusion constructs imaged both in-vitro and on the plasma membrane of HeLa cells. We present an application of these methods to the imaging of a pair of plasma membrane proteins involved in cell signaling.

[1] P. Annibale et al. Nature Methods 8 (2011) 527-528