Pince optique et microscopie à contraste de phase pour l'étude de la mécanique cellulaire : développement, modélisation et calibration en réflexion.

Flavie Gillant 1
1 Laboratoire Charles Fabry / Biophotonique
LCF - Laboratoire Charles Fabry
Abstract : This manuscript details the development of an optical tweezer setup to study the mechanical properties of endothelial cells, involved in the development of atherosclerosis. The goal is to determine the viscoelastic properties of the cells, and to follow the propagation of the mechanical constraint inside the cell. This mechanical constraint is applied via a bead attached to the cell membrane and subjected to an optical trap.The setup built combines optical trapping with phase contrast microscopy, to apply a force while imaging the cells with the same microscope objective. The originality of the optical tweezer setup relies on the detection of the signal backscattered by the trapped bead, in a plane conjugate to the back focal plane of the objective, in order to measure the relative position of the bead with respect to the center of the trap.An important part of this work was dedicated to the understanding of the detected signal presenting an interference pattern with rings, explained by a simple model. This model provides an explanation for the position measurement artifacts arising from the superposition of the phase ring and the interference pattern. To solve the problem, the phase ring was moved in a conjugate plane involved only in the imaging path of the sample.The interference pattern has the main advantage of giving access to the precise height of the trapped bead, usually difficult to measure. This information is necessary to calibrate the optical trap stiffness at the height of the cells, either by the power spectrum analysis of the Brownian motion of the trapped bead, or by its response to a step motion of the trap. These two calibration methods, along with the application of the equipartition theorem and Bayesian inference analysis, were implemented and their results compared, showing a good agreement. The complete calibration of the setup makes it a ready-to-use tool to exert local forces controlled in direction and amplitude on cells.
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Flavie Gillant. Pince optique et microscopie à contraste de phase pour l'étude de la mécanique cellulaire : développement, modélisation et calibration en réflexion.. Optique [physics.optics]. Université Paris-Saclay, 2016. Français. ⟨NNT : 2016SACLO016⟩. ⟨tel-01629471⟩

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