Brillouin Scattering Microscopy for Mechanical Imaging

Abstract

In a world where science is constantly challenged to solve problems of increasing complexity, light is paving new ways to gather information about the physical properties of matter. Among these properties, elasticity is becoming fundamental in the understanding and the diagnosis of several diseases. Current solutions to gather mechanical information, however, measure the response of a material to an applied excitation, which makes them invasive and limited by a low spatial resolution. In contrast with these techniques, Brillouin spectroscopy offers the unique solution to retrieve stiffness information from the spectrum of the light scattered by inherent thermal acoustic waves. The combination of Brillouin spectroscopy with confocal microscopy has yielded a confocal Brillouin microscope able to perform mechanical imaging in a non-invasive manner. This was used to investigate two different biological problems: on the one hand the stiffness variations in specific endothelium cells of the eye, aiming at a better understanding of the mechanisms responsible for glaucoma, and on the other the characterisation of the mechanical structures of blood vessels, which could provide fundamental information regarding the formation of atherosclerotic plaques. Following an investigation on the optimal geometry that minimises the spectral broadening caused by the collection of photons over a range of scattering angles, high resolution Brillouin imaging was obtained in a confocal backscattering arrangement. To the best of our knowledge this thesis presents, for the first time, sub-cellular Brillouin images. In particular, in vitro Brillouin images of single HUVEC cells were acquired to investigate the cell’s mechanical response to the application of the Latrunculin-A drug. This analysis, together with the finding of a linear correlation between the Brillouin modulus and the standard Young’s modulus, validates the technique as a feasible means of measuring stiffness. Following this assessment, Brillouin images of normal and diseased vessels were acquired showing that the atherosclerotic plaques had a lower stiffness compared to both diseased and healthy vessel walls. These results might encourage the application of confocal Brillouin microscopy as the tool of choice for the investigation of the arterial biomechanics.