Optimizing 1-渭s-Resolution Single-Molecule Force Spectroscopy on a Commercial Atomic Force Microscope

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• 作者：Devin T. Edwards ; Jaevyn K. Faulk ; Aric W. Sanders ; Matthew S. Bull ; Robert Walder ; Marc-Andre LeBlanc ; Marcelo C. Sousa ; Thomas T. Perkins
• 刊名：Nano Letters
• 出版年：2015
• 出版时间：October 14, 2015
• 年：2015
• 卷：15
• 期：10
• 页码：7091-7098
• 全文大小：450K
• ISSN：1530-6992

文摘

Atomic force microscopy (AFM)-based single-molecule force spectroscopy (SMFS) is widely used to mechanically measure the folding and unfolding of proteins. However, the temporal resolution of a standard commercial cantilever is 50鈥?000 渭s, masking rapid transitions and short-lived intermediates. Recently, SMFS with 0.7-渭s temporal resolution was achieved using an ultrashort (L = 9 渭m) cantilever on a custom-built, high-speed AFM. By micromachining such cantilevers with a focused ion beam, we optimized them for SMFS rather than tapping-mode imaging. To enhance usability and throughput, we detected the modified cantilevers on a commercial AFM retrofitted with a detection laser system featuring a 3-渭m circular spot size. Moreover, individual cantilevers were reused over multiple days. The improved capabilities of the modified cantilevers for SMFS were showcased by unfolding a polyprotein, a popular biophysical assay. Specifically, these cantilevers maintained a 1-渭s response time while eliminating cantilever ringing (Q 鈮?0.5). We therefore expect such cantilevers, along with the instrumentational improvements to detect them on a commercial AFM, to accelerate high-precision AFM-based SMFS studies.