Cellular Force Microscope.

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Over the past decade the field of mechanobiology has expanded exponentially due to a growing realisation that the coupling between mechanical properties and biochemistry plays a central role in biology and potentially in a number of diseases. This expansion has been enabled by the adoption of a number of techniques for measuring mechanical properties at a cellular and sub-cellular scale, arguably the most widely used of which is atomic force microscopy (AFM). By indenting a sharp probe attached to a force sensing cantilever into a sample surface, a curve of indentation against force can be measured, and from this the sample's elastic modulus can be calculated. AFM instrument manufacturers have made the process relatively straightforward, automated the analysis, and have spawned the growth of an active scientific community. The numbers obtained are now used to inform models and improve understanding of fundamental biological processes such as cellular motility, endocytosis and tissue development, as well as to drive translational aspects such as tissue repair and intervention. Unfortunately there is a fundamental problem in that the measurements of modulus obtained contain systematic errors typically of 10s to 100s of percent - the force sensor, the AFM cantilever, is simply not sensitive enough to accurately measure the small forces asked of it.To address this problem we will use a new generation of small and soft AFM cantilevers that have the required sensitivity, we will build a detection system that enables accurate measurement of their stiffness and deflection, removing another major source of measurement error, and house it within a custom designed instrument able to measure indentation accurately over 10s of micrometres. The resultant Cellular Force Microscope (CFM) will have the sensitivity and accuracy necessary to measure the properties of soft cells and tissue.We will use the instrument to look at two example systems in which we have considerable experience. Nerves in the peripheral nervous system (PNS) contain a number of different cell types, including neurones and Schwann cells, both of which are exceptionally soft even for animal cells. Traumatic damage to the PNS is common and can result in loss of feeling and function. Nerve repair remains a considerable challenge, and better understanding of nerve mechanical properties has an important role to play in the selection of suitable tissue engineering scaffolds for nerve regeneration. We will use our new instrument to obtain preliminary data on the mechanical properties of cells from the PNS, with the aim of informing the selection of mechanical property matching materials. The second system we will study is the mechanics of the soft tissue in the interior of bone. This is tissue that is mechanically heterogeneous but contains regions that are very soft and as such is an excellent example of the kind of advanced, more in vivo, mechanobiology study that AFM is starting to be used for. As part of a project on breast cancer metastasis we are currently characterising the properties of bone using conventional AFM so it provides an excellent tissue model for applying the new technology, using surplus bone tissue. Our new data will contribute to that work on understanding whether the mechanics of the bone influences the secondary spread of cancer to this site.