A Quick and Reproducible Silanization Method by Using Plasma Activation for Hydrophobicity-Based Kinesin Single Molecule Fluorescence–Microscopy Assays |
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Authors: | Viktoria Wedler Dustin Quinones Prof?Dr Heiko Peisert Prof?Dr Erik Schäffer |
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Institution: | 1. Eberhard Karls Universität Tübingen, Cellular Nanoscience (ZMBP), Auf der Morgenstelle 32, 72076 Tübingen, Germany;2. Eberhard Karls Universität Tübingen, Institute of Physical and Theoretical Chemistry, Auf der Morgenstelle 18, 72076 Tübingen, Germany |
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Abstract: | Single-molecule assays often require functionalized surfaces. One approach for microtubule assays renders surfaces hydrophobic and uses amphiphilic blocking agents. However, the optimal hydrophobicity is unclear, protocols take long, produce toxic waste, and are susceptible to failure. Our method uses plasma activation with hydrocarbons for hexamethyldisilazane (HMDS) silanization in the gas phase. We measured the surface hydrophobicity, its effect on how well microtubule filaments were bound to the surface, and the number of nonspecific interactions with kinesin motor proteins. Additionally, we tested and discuss the use of different silanes and activation methods. We found that even weakly hydrophobic surfaces were optimal. Our environmentally friendly method significanty reduced the overall preparation effort and resulted in reproducible, high-quality surfaces with low variability. We expect the method to be applicable to a wide range of other single-molecule assays. |
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Keywords: | Surface chemistry single-molecule fluorescence silanes photoelectron spectroscopy microtubules |
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