Improved single molecule force spectroscopy using micromachined cantilevers

Enhancing the short-term force precision of atomic force microscopy (AFM) while maintaining excellent long-term force stability would result in improved performance across multiple AFM modalities, including single molecule force spectroscopy (SMFS). SMFS is a powerful method to probe the nanometer-scale dynamics and energetics of biomolecules (DNA, RNA, and proteins). The folding and unfolding rates of such macromolecules are sensitive to sub-pN changes in force. Recently, we demonstrated sub-pN stability over a broad bandwidth (Δf = 0.01–16 Hz) by removing the gold coating from a 100 μm long cantilever. However, this stability came at the cost of increased short-term force noise, decreased temporal response, and poor sensitivity. Here, we avoided these compromises while retaining excellent force stability by modifying a short (L = 40 μm) cantilever with a focused ion beam. Our process led to a ∼10-fold reduction in both a cantilever’s stiffness and its hydrodynamic drag near a surface. We also preserved ...

[1]  Julio M Fernandez,et al.  Force-Clamp Spectroscopy Monitors the Folding Trajectory of a Single Protein , 2004, Science.

[2]  Lucille A. Giannuzzi,et al.  Introduction to Focused Ion Beams , 2005 .

[3]  David Baker,et al.  Computer-based redesign of a protein folding pathway , 2001, Nature Structural Biology.

[4]  J. Latgé,et al.  High-resolution imaging of chemical and biological sites on living cells using peak force tapping atomic force microscopy. , 2012, Langmuir : the ACS journal of surfaces and colloids.

[5]  Evan Evans,et al.  Five challenges to bringing single-molecule force spectroscopy into living cells , 2011, Nature Methods.

[6]  C. W. Hagen,et al.  A critical literature review of focused electron beam induced deposition , 2008 .

[7]  J. Liphardt,et al.  Reversible Unfolding of Single RNA Molecules by Mechanical Force , 2001, Science.

[8]  Andrew C. Richardson,et al.  Quantifying noise in optical tweezers by allan variance. , 2009, Optics express.

[9]  Toshio Ando,et al.  High-speed atomic force microscopy coming of age , 2012, Nanotechnology.

[10]  Daniel J Müller,et al.  Atomic force microscopy: a nanoscopic window on the cell surface. , 2011, Trends in cell biology.

[11]  Daniel J. Muller,et al.  Imaging and quantifying chemical and physical properties of native proteins at molecular resolution by force-volume AFM. , 2011, Angewandte Chemie.

[12]  Runcong Liu,et al.  Correction of the viscous drag induced errors in macromolecular manipulation experiments using atomic force microscope. , 2010, Review of Scientific Instruments.

[13]  Matthias Rief,et al.  Ligand-Dependent Equilibrium Fluctuations of Single Calmodulin Molecules , 2009, Science.

[14]  V. Dötsch,et al.  Single-molecule force spectroscopy from nanodiscs: an assay to quantify folding, stability, and interactions of native membrane proteins. , 2012, ACS nano.

[15]  T. Perkins,et al.  Routine and timely sub-picoNewton force stability and precision for biological applications of atomic force microscopy. , 2012, Nano letters.

[16]  S. Scheuring,et al.  Automated setpoint adjustment for biological contact mode atomic force microscopy imaging , 2010, Nanotechnology.

[17]  Hongbin Li,et al.  Direct observation of markovian behavior of the mechanical unfolding of individual proteins. , 2008, Biophysical journal.

[18]  Carlos Bustamante,et al.  Direct Observation of the Three-State Folding of a Single Protein Molecule , 2005, Science.

[19]  G. Neuert,et al.  Thiol-based, site-specific and covalent immobilization of biomolecules for single-molecule experiments , 2010, Nature Protocols.

[20]  Simon Scheuring,et al.  Chromatic Adaptation of Photosynthetic Membranes , 2005, Science.

[21]  Julie Gold,et al.  Protein Adsorption on Model Surfaces with Controlled Nanotopography and Chemistry , 2002 .

[22]  Ashley R. Carter,et al.  Ultrastable atomic force microscopy: atomic-scale stability and registration in ambient conditions. , 2009, Nano letters.

[23]  Daniel J Müller,et al.  Atomic force microscopy as a multifunctional molecular toolbox in nanobiotechnology. , 2008, Nature nanotechnology.

[24]  Taekjip Ha,et al.  Initiation and re-initiation of DNA unwinding by the Escherichia coli Rep helicase , 2002, Nature.

[25]  Daniel J Müller,et al.  Force probing surfaces of living cells to molecular resolution. , 2009, Nature chemical biology.

[26]  A. Engel,et al.  Structural changes in native membrane proteins monitored at subnanometer resolution with the atomic force microscope: a review. , 1997, Journal of structural biology.

[27]  M. Rief,et al.  Reversible unfolding of individual titin immunoglobulin domains by AFM. , 1997, Science.

[28]  Manel Puig-Vidal,et al.  High-Speed Force Spectroscopy Unfolds Titin at the Velocity of Molecular Dynamics Simulations , 2013, Science.

[29]  Andreas Engel,et al.  Structure and mechanics of membrane proteins. , 2008, Annual review of biochemistry.

[30]  P. Grütter,et al.  The noise of coated cantilevers , 2012, Nanotechnology.

[31]  T. Vystavěl,et al.  Focused ion beam (FIB) milling of electrically insulating specimens using simultaneous primary electron and ion beam irradiation , 2007 .

[32]  Chanmin Su,et al.  Mechanical mapping of single membrane proteins at submolecular resolution. , 2011, Nano letters.

[33]  Matthias Rief,et al.  Single Molecule Force Spectroscopy on Polysaccharides by Atomic Force Microscopy , 1997, Science.

[34]  Xu,et al.  "Dip-Pen" nanolithography , 1999, Science.

[35]  Markus Pollnau,et al.  Focused ion beam scan routine, dwell time and dose optimizations for submicrometre period planar photonic crystal components and stamps in silicon , 2007 .

[36]  H. Gaub,et al.  Unfolding forces of titin and fibronectin domains directly measured by AFM. , 2000, Advances in experimental medicine and biology.

[37]  M. Woodside,et al.  Direct observation of multiple misfolding pathways in a single prion protein molecule , 2012, Proceedings of the National Academy of Sciences.

[38]  Nabil A. Amro,et al.  Production of Nanostructures of DNA on Surfaces , 2002 .

[39]  Charles M. Lieber,et al.  Stretching and breaking duplex DNA by chemical force microscopy. , 1997, Chemistry & biology.

[40]  Lucille A. Giannuzzi,et al.  Introduction to focused ion beams : instrumentation, theory, techniques, and practice , 2010 .

[41]  A. Engel,et al.  Electrostatically balanced subnanometer imaging of biological specimens by atomic force microscope. , 1999, Biophysical journal.

[42]  Jason Cleveland,et al.  Finite optical spot size and position corrections in thermal spring constant calibration , 2004 .

[43]  K. Svoboda,et al.  Biological applications of optical forces. , 1994, Annual review of biophysics and biomolecular structure.

[44]  J. Howard,et al.  Mechanics of Motor Proteins and the Cytoskeleton , 2001 .

[45]  D. Mariolle,et al.  Reduction of the cantilever hydrodynamic damping near a surface by ion-beam milling , 2006 .

[46]  D. Müller,et al.  Multiparametric imaging of biological systems by force-distance curve–based AFM , 2013, Nature Methods.

[47]  G Büldt,et al.  Imaging purple membranes in aqueous solutions at sub-nanometer resolution by atomic force microscopy. , 1995, Biophysical journal.

[48]  Daniel J Müller,et al.  Single-molecule studies of membrane proteins. , 2006, Current opinion in structural biology.

[49]  M. Viani,et al.  Small cantilevers for force spectroscopy of single molecules , 1999 .

[50]  Hongbin Li,et al.  A functional single-molecule binding assay via force spectroscopy , 2007, Proceedings of the National Academy of Sciences.

[51]  N. Vieira,et al.  Imaging carious human dental tissue with optical coherence tomography , 2006 .

[52]  Alex R. Hodges,et al.  Improved atomic force microscope cantilever performance by ion beam modification , 2001 .

[53]  T. Perkins,et al.  Atomic force microscopy with sub-picoNewton force stability for biological applications. , 2013, Methods.

[54]  M. Rief,et al.  Sequence-dependent mechanics of single DNA molecules , 1999, Nature Structural Biology.

[55]  Daniel J. Muller,et al.  Characterizing molecular interactions in different bacteriorhodopsin assemblies by single-molecule force spectroscopy. , 2006, Journal of molecular biology.

[56]  M. Rief,et al.  The Complex Folding Network of Single Calmodulin Molecules , 2011, Science.

[57]  A. Engel,et al.  Native Escherichia coli OmpF porin surfaces probed by atomic force microscopy. , 1995, Science.

[58]  Jens Struckmeier,et al.  Hydrodynamic effects in fast AFM single-molecule force measurements , 2005, European Biophysics Journal.