Sweers et al. Nanoscale Research Letters 2011, 6:270 http://www.nanoscalereslett.com/content/6/1/270 N A N O E X P R E S S Open Access Nanomechanical properties of a-synuclein amyloid fibrils: a comparative study by nanoindentation, harmonic force microscopy, and Peakforce QNM Kim Sweers*, Kees van der Werf, Martin Bennink and Vinod Subramaniam* Abstract We report on the use of three different atomic force spectroscopy modalities to determine the nanomechanical properties of amyloid fibrils of the human a-synuclein protein. a-Synuclein forms fibrillar nanostructures of approximately 10 nm diameter and lengths ranging from 100 nm to several microns, which have been associated with Parkinson’s disease. Atomic force microscopy (AFM) has been used to image the morphology of these protein fibrils deposited on a flat surface. For nanomechanical measurements, we used single-point nanoindentation, in which the AFM tip as the indenter is moved vertically to the fibril surface and back while the force is being recorded. We also used two recently developed AFM surface property mapping techniques: Harmonic force microscopy (HarmoniX) and Peakforce QNM. These modalities allow extraction of mechanical parameters of the surface with a lateral resolution and speed comparable to tapping-mode AFM imaging. Based on this phenomenological study, the elastic moduli of the a-synuclein fibrils determined using these three different modalities are within the range 1.3-2.1 GPa. We discuss the relative merits of these three methods for the determination of the elastic properties of protein fibrils, particularly considering the differences and difficulties of each method. Introduction Amyloid fibrils are insoluble protein aggregates that have been associated with a range of neurodegenerative diseases, including Huntington, Alzheimer’s, Parkinson’s, and Creutzfeldt-Jakob disease [1]. The fibrils typically have a diameter ranging from 4 to 12 nm, and lengths from 100 nm up to several microns [2-4]. In this study, we investigated the nanomechanical properties of amy- loid fibrils formed from the human a-synuclein protein, which is associated with Parkinson’s disease. a-Synuclein amyloid fibrils are found in the brains of Parkinson’s disease patients as components of larger plaques called Lewy bodies [5,6]. Atomic force microscopy (AFM) has been primarily used as an imaging tool to determine morphological parameters such as height and length of amyloid fibrils, * Correspondence: k.k.m.sweers@utwente.nl; v.subramaniam@utwente.nl Nanobiophysics Group, MESA+ Institute for Nanotechnology, Faculty of Science and Technology, University of Twente, Enschede, The Netherlands such as those formed from a-synuclein [2-4], insulin [7], and b-lactoglobulin [8]. AFM is also a powerful techni- que for characterizing mechanical properties. With the ability to exert and measure forces up to the piconewton range, AFM is a particularly suitable tool to determine the nanomechanical properties of nanometer-sized bio- logical structures, such as amyloid fibrils. Mechanical properties such as stiffness, rigidity, resistance to break- age or adhesive properties of these fibrils or individual monomers are interesting for the use of these fibrils as nanomaterials, for getting a better understanding of the physico-chemical properties of these fibrils, and to get more insight into their structure and growth [9-14]. Indentation-type AFM or single-point nanoindentation (SPI), for example, implemented as ‘Point-and-Shoot’ in the Veeco operating software, is the most widely used method to measure nanomechanical properties of a sample. In this mode, the tip approaches and indents the sample until a certain predefined force is reached. © 2011 Sweers et al; licensee Springer. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.