A Quick and Reproducible Silanization Method by Using Plasma Activation for Hydrophobicity‐Based Kinesin Single Molecule Fluorescence–Microscopy Assays. Issue 64 (14th September 2022)

• Record Type: Journal Article
• Authors: Wedler, Viktoria; Quinones, Dustin; Peisert, Heiko; Schäffer, Erik
• Is Part Of: Chemistry. Volume 28:Issue 64(2022)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2022-09-14
• Subjects: Surface chemistry -- single-molecule fluorescence -- silanes -- photoelectron spectroscopy -- microtubules; Chemistry -- Periodicals; 540
• DOI: 10.1002/chem.202202036
• Languages: English
• ISSNs: 0947-6539

Abstract

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. Abstract : Strength in weakness : Weakly hydrophobic, plasma‐activated surfaces silanized with a disilazane turned out to be optimal for single‐molecule fluorescence measurements of biological molecular machines. The novel silanization assay reduced the work load more than tenfold and does not require any organic solvents, thus making the assay environmentally friendly and efficient.