title: Nanomechanical assessment of human and murine collagen fibrils via atomic force microscopy cantilever-based nanoindentation.
abstract: The nanomechanical assessment of collagen fibrils via atomic force microscopy (AFM) is of increasing interest within the biomedical research community. In contrast to conventional nanoindentation there exists no common standard for conducting experiments and analysis of data. Currently used analysis approaches vary between studies and validation of quantitative results is usually not performed, which makes comparison of data from different studies difficult. Also there are no recommendations with regards to the maximum indentation depth that should not be exceeded to avoid substrate effects. Here we present a methodology and analysis approach for AFM cantilever-based nanoindentation experiments that allows efficient use of captured data and relying on a reference sample for determination of tip shape. Further we show experimental evidence that maximum indentation depth on collagen fibrils should be lower than 10-15% of the height of the fibril to avoid substrate effects and we show comparisons between our and other approaches used in previous works. While our analysis approach yields similar values for indentation modulus compared to the Oliver-Pharr method we found that Hertzian analysis yielded significantly lower values. Applying our approach we successfully and efficiently indented collagen fibrils from human bronchi, which were about 30 nm in size, considerably smaller compared to collagen fibrils obtained from murine tail-tendon. In addition, derived mechanical parameters of collagen fibrils are in agreement with data previously published. To establish a quantitative validation we compared indentation results from conventional and AFM cantilever-based nanoindentation on polymeric samples with known mechanical properties. Importantly we can show that our approach yields similar results when compared to conventional nanoindentation on polymer samples. Introducing an approach that is reliable, efficient and taking into account the AFM tip shape, we anticipate that the present work may act as a guideline for conducting AFM cantilever-based nanoindentation of collagen fibrils. This may aid understanding of collagen-related diseases such as asthma, lung fibrosis or bone disease with potential alterations of collagen fibril mechanics.
authors.affiliation: Bioengineering Science Research Group, Faculty of Engineering and the Environment, University of Southampton, SO17 1BJ Southampton, UK; Institute for Lightweight Design and Structural Biomechanics, Faculty of Engineering and the Environment, Vienna University of Technology, Gusshausstrasse 27-29, Vienna 1040, Austria. Electronic address: oandriot@ilsb.tuwien.ac.at.
authors.forename: Orestis G
authors.initials: OG
authors.lastname: Andriotis
authors.affiliation: The Brooke Laboratories, Division of Infection, Inflammation and Immunity, Faculty of Medicine, University of Southampton, SO16 6YD, Southampton, UK.
authors.forename: Wiparat
authors.initials: W
authors.lastname: Manuyakorn
authors.affiliation: National Centre of Advanced Tribology at Southampton, Faculty of Engineering and the Environment, University of Southampton, SO17 1BJ Southampton, UK.
authors.forename: Jurgita
authors.initials: J
authors.lastname: Zekonyte
authors.affiliation: Bioengineering Science Research Group, Faculty of Engineering and the Environment, University of Southampton, SO17 1BJ Southampton, UK.
authors.forename: Orestis L
authors.initials: OL
authors.lastname: Katsamenis
authors.affiliation: Bioengineering Science Research Group, Faculty of Engineering and the Environment, University of Southampton, SO17 1BJ Southampton, UK.
authors.forename: Sebastien
authors.initials: S
authors.lastname: Fabri
authors.affiliation: The Brooke Laboratories, Division of Infection, Inflammation and Immunity, Faculty of Medicine, University of Southampton, SO16 6YD, Southampton, UK.
authors.forename: Peter H
authors.initials: PH
authors.lastname: Howarth
authors.affiliation: The Brooke Laboratories, Division of Infection, Inflammation and Immunity, Faculty of Medicine, University of Southampton, SO16 6YD, Southampton, UK.
authors.forename: Donna E
authors.initials: DE
authors.lastname: Davies
authors.affiliation: Bioengineering Science Research Group, Faculty of Engineering and the Environment, University of Southampton, SO17 1BJ Southampton, UK; Institute for Lightweight Design and Structural Biomechanics, Faculty of Engineering and the Environment, Vienna University of Technology, Gusshausstrasse 27-29, Vienna 1040, Austria. Electronic address: pthurner@ilsb.tuwien.ac.at.
authors.forename: Philipp J
authors.initials: PJ
authors.lastname: Thurner
mesh_terms: D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry*; D001696:Biomechanical Phenomena; D001980:Bronchi / Q000473:pathology; D003094:Collagen / Q000737:chemistry*; D004548:Elasticity; D006801:Humans; D008422:Materials Testing; D051379:Mice; D018625:Microscopy, Atomic Force; D049329:Nanostructures / Q000737:chemistry*; D036103:Nanotechnology / Q000379:methods*; D016012:Poisson Distribution; D011108:Polymers / Q000737:chemistry; D051381:Rats; D015203:Reproducibility of Results; D013314:Stress, Mechanical
chemical_list: D001672:Biocompatible Materials; D011108:Polymers; D003094:Collagen
keywords: Airways; Atomic force microscopy; Bronchi; Collagen; Indentation; Mechanics; Mouse; Rat