Elastic-resonator-interference-stress-microscopy (ERISM)
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Date
23/06/2017Author
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Abstract
The forces biological cells apply to their environment are recognised to be critical during
processes like migration, division, wound healing, and stem cell differentiation. Methods to
measure these forces have been extremely valuable in contributing to our understanding of
cell-substrate and cell-cell interactions. However, existing force sensing techniques struggle
to measure forces cells apply perpendicular to the plane of their substrate although these
out-of-plane forces have been demonstrated to be important in many processes. In addition,
most currently used force sensing techniques require fluorescence imaging which can lead to
photo-toxic effects if high frame rates are required. Finally, many methods require detaching
of cells after the measurement which prevents measuring the same cells repeatedly or using
immunostaining, which is an important tool for linking biomechanical and biochemical
observations.
In this thesis, we introduce a novel high-throughput and low-light-intensity force sensing
technique which is inherently well suited to measure vertical forces. Elastic-Resonator-Interference-Stress-Microscopy (ERISM) measures the spatially resolved reflectance of an
elastic micro-cavity. With fully automated hyperspectral imaging and data analysis supported
by transfer-matrix modelling, this allows tracking of nanometre thickness changes
across a large area of the cavity. By combining Atomic-Force-Microscopy with a Finite-Element-Method,
we extract basic material properties of the micro-cavities to calculate
stress and force.
Using the example of different neural cells, we provide experimental evidence that ERISM
measurements can be performed over hours at high frame rates or repeatedly over weeks
for the same sample to investigate a variety of cellular processes like cell spreading, growth cone migration, and stem cell differentiation. We perform immunostaining for cell specific
markers on ERISM micro-cavities as detaching of the cells is not required. Furthermore, we
find that the high throughput of ERISM allows us to find significant differences between
wild-type and knock-out cell populations for a gene associated with dyslexia.
Type
Thesis, PhD Doctor of Philosophy
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