TY - JOUR
KW - Structural Biology
AU - Devin Edwards
AU - Thomas Perkins
AB - Atomic force microscopy (AFM)-based single-molecule force spectroscopy (SMFS) enables a wide array of studies, from measuring the strength of a ligand-receptor bond to elucidating the complex folding pathway of individual membrane proteins. Such SMFS studies and, more generally, the diverse applications of AFM across biophysics and nanotechnology are improved by enhancing data quality via improved force stability, force precision, and temporal resolution. For an advanced, small-format commercial AFM, we illustrate how these three metrics are limited by the cantilever itself rather than the larger microscope structure, and then describe three increasingly sophisticated cantilever modifications that yield enhanced data quality. First, sub-pN force precision and stability over a broad bandwidth (Delta f = 0.01-20 Hz) is routinely achieved by removing a long (L = 100 mu m) cantilever's gold coating. Next, this sub-pN bandwidth is extended by a factor of similar to 50 to span five decades of bandwidth (Delta f = 0.01-1000 Hz) by using a focused ion beam (FIB) to modify a shorter (L = 40 mu m) cantilever. Finally, FIB-modifying an ultrashort (L = 9 mu m) cantilever improves its force stability and precision while maintaining 1-mu s temporal resolution. These modified ultrashort cantilevers have a reduced quality factor Q approximate to 0.5) and therefore do not apply a substantial (30-90 pN), high-frequency force modulation to the molecule, a phenomenon that is unaccounted for in traditional SMFS analysis. Currently, there is no perfect cantilever for all applications. Optimizing AFM-based SMFS requires understanding the tradeoffs inherent to using a specific cantilever and choosing the one best suited to a particular application. Published by Elsevier Inc.
BT - Journal of Structural Biology
DA - 2017-01
DO - 10.1016/j.jsb.2016.01.009
IS - 1
N2 - Atomic force microscopy (AFM)-based single-molecule force spectroscopy (SMFS) enables a wide array of studies, from measuring the strength of a ligand-receptor bond to elucidating the complex folding pathway of individual membrane proteins. Such SMFS studies and, more generally, the diverse applications of AFM across biophysics and nanotechnology are improved by enhancing data quality via improved force stability, force precision, and temporal resolution. For an advanced, small-format commercial AFM, we illustrate how these three metrics are limited by the cantilever itself rather than the larger microscope structure, and then describe three increasingly sophisticated cantilever modifications that yield enhanced data quality. First, sub-pN force precision and stability over a broad bandwidth (Delta f = 0.01-20 Hz) is routinely achieved by removing a long (L = 100 mu m) cantilever's gold coating. Next, this sub-pN bandwidth is extended by a factor of similar to 50 to span five decades of bandwidth (Delta f = 0.01-1000 Hz) by using a focused ion beam (FIB) to modify a shorter (L = 40 mu m) cantilever. Finally, FIB-modifying an ultrashort (L = 9 mu m) cantilever improves its force stability and precision while maintaining 1-mu s temporal resolution. These modified ultrashort cantilevers have a reduced quality factor Q approximate to 0.5) and therefore do not apply a substantial (30-90 pN), high-frequency force modulation to the molecule, a phenomenon that is unaccounted for in traditional SMFS analysis. Currently, there is no perfect cantilever for all applications. Optimizing AFM-based SMFS requires understanding the tradeoffs inherent to using a specific cantilever and choosing the one best suited to a particular application. Published by Elsevier Inc.
PB - Elsevier BV
PY - 2017
SP - 13
EP - 25
T2 - Journal of Structural Biology
TI - Optimizing force spectroscopy by modifying commercial cantilevers: Improved stability, precision, and temporal resolution
VL - 197
SN - 1047-8477
ER -