Dresden 2011 – scientific programme

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BP: Fachverband Biologische Physik

BP 30: Posters: Physics of Cells

BP 30.11: Poster

Thursday, March 17, 2011, 17:15–20:00, P3

Quantitative TIRF Microscopy of Fluorescent Layers — •Haugen Grefe and Hans-Günther Döbereiner — Institut für Biophysik, Universität Bremen, Germany

Lamellipodia play an important role for the motility of cells. Our aim is to measure the growth dynamics and thickness of these structures using total internal reflection fluorescence (TIRF) microscopy. Upon sending a laser beam on a cover slip with an angle above the critical angle of total reflection an evanescent intensity field appears in the preparation behind the glass. The penetration depth is in the range of 50 to 1000 nm, which is also the expected thickness of lamellipodia. When one linearly increases the laser angle the fluorescence intensity of excited fluorophores behind the cover slip decreases exponentially. Fitting the intensity as a function of penetration depth gives the size of the fluorescent object. This procedure works well with dyed latex beads with a diameter of 100 to 500 nm. In the next step we will produce fluorescent layers with a defined variable thickness. The intention is to get a scale for measuring dyed lamellipodia as well as to optimize the theoretical background for fitting the intensity result.