Application of Large Area Mapping AFM for Automated Structural and Mechanical Analysis of Developing Cells and Tissues

Session

Imaging Biomechanics

Authors

Dr Alexander Dulebo (1), Dr. Tanja Neumann (1), Dr. André Körnig (1), Dr. Torsten Müller (1), Dr.-Ing. Dimitar Stamov (1), Dr. Heiko Haschke (1)

Affiliations

1. JPK BioAFM, Bruker Nano GmbH

Keywords

Biomechanics, Mechanobiology, Cytoskeleton Imaging, Life Cell Imaging, AFM, Imaging - advanced

Abstract text

AFM can be successfully applied for comprehensive nano-mechanical characterization of single molecules, cells and tissues, under near physiological conditions [1]. Currently, the trend is to extend this by studying the mechanobiology of living cells while evaluating their structure and the interaction with their cell culture substrates [2,3]. It is interesting to understand how cell behaviour is driven by the cytoskeletal dynamics and cell mechanics in typical cell culture scaffold scenarios. We will introduce the concept of automated large area multiparametric characterization of densely packed cell layers and highly corrugated tissue samples.

We applied high-speed AFM, with a temporal resolution on the second to millisecond scale to resolve dynamic processes such as the collagen fibrillogenesis and cytoskeletal dynamics in living cells. As a tool for analyzing the complex cellular mechanobiology, we went beyond purely elastic models, and performed sine oscillations (up to 1 kHz, amplitude 5-60 nm) in Z while in contact with the surface to probe the frequency-dependent response of living fibroblasts.

We will provide insight into the structural formation of collagen type I, emphasizing the intermediate steps in the process [4]. We will demonstrate how cell spreading and migration in living KPG-7 fibroblasts and CHO cells, can be associated with spatially resolved cytoskeletal reorganization events. We will further discuss how to calculate the viscoelastic properties, characterized by the dynamic storage and loss modulus (E’, E’’) distribution in living fibroblast cells. In the past, investigating large and rough samples such as tissues and hydrogels using AFM was challenging due to the limited z-axis of the AFM. Using osteoarthritic cartilage as an example, we will demonstrate how a newly developed stage enables multi-region AFM probing over a large, rough sample area while providing additional correlative optical data sets [5].

We will discuss how these developments, in combination with advanced optical microscopy techniques, can overcome the inherent drawbacks of traditional AFM systems for characterizing challenging biological samples.

References

[1] M.W Amrein, and D. Stamov, D (2019). Atomic Force Microscopy in the Life Sciences in Springer Handbook of Microscopy (eds P.W. Hawkes and J.C.H. Spence), Springer International Publishing, pp 1469–1505.
[2] P. Elter, T. Weihe, R. Lange, et al (2011) Eur Biophys J 40(3):317–327.
[3] A.J. Engler, S. Sen, H.L. Sweeney, and D.E. Discher (2006) Cell 126(4):677–689.
[4] D.R. Stamov, E. Stock, C.M. Franz, et al (2015), Ultramicroscopy 149:86-94.
[5] M. Tschaikowsky, T. Neumann, S. Brander, et al (2021), Acta Biomater 126:315-325.