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Nano-Scale Dynamic Recognition Imaging on Vascular Endothelial Cells

Lilia A. Chtcheglova ^*, Jens Waschke , Linda Wildling ^*, Detlev Drenckhahn  and Peter Hinterdorfer ^*

^* Institute for Biophysics, Johannes Kepler University of Linz, Linz, Austria; and Institute of Anatomy and Cell Biology, University of Würzburg, Würzburg, Germany

Correspondence: Address reprint requests and inquiries to P. Hinterdorfer, Tel.: 43-732-2468-9265; E-mail: peter.hinterdorfer{at}jku.at.

Combination of high-resolution atomic force microscope topography imaging with single molecule force spectroscopy provides a unique possibility for the detection of specific molecular recognition events. The identification and localization of specific receptor binding sites on complex heterogeneous biosurfaces such as cells and membranes are of particular interest in this context. Here simultaneous topography and recognition imaging (TREC) was applied to gently fixed microvascular endothelial cells from mouse myocardium (MyEnd) to identify binding sites of vascular endothelial (VE)-cadherin, known to play a crucial role in calcium-dependent, homophilic cell-to-cell adhesion. TREC images were acquired with magnetically oscillating atomic-force microscope tips functionalized with a recombinant VE-cadherin-Fc cis-dimer. The recognition images revealed single molecular binding sites and prominent, irregularly shaped dark spots (domains) with sizes ranging from 10 to 100 nm. These domains arose from a decrease of the oscillation amplitude during specific binding between active VE-cadherin cis-dimers. The VE-cadherin clusters were subsequently assigned to topography features. TREC represents an exquisite method to quickly obtain the local distribution of receptors on cellular surface with an unprecedented lateral resolution of 5 nm.

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