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Microscopie par génération de troisième harmonique appliquée à la biologie.

Abstract : This work aimed at developing third-harmonic generation microscopy for biology, a recent technique allowing the visualization of unstained cells and tissues with sub-micrometer resolution. Its application to biological imaging was to date limited by the lack of study of signal creation inside samples, endogenous sources of contrast in biology, and induced phototoxicity. The work presented here mainly addresses those three issues. Firstly, we studied theoretically and experimentally the influence of sample structure and focusing of the excitation beam on THG signal. We showed that THG imaging acts on the sample as a bandpass filter for spatial frequencies and that adjusting the focusing of the excitation beam allows to modulate the signal from structures different shapes inside a complex sample. Secondly, we characterized optical properties of various biologically relevant liquids, showing that lipid bodies embedded in an aqueous environment should provide a strong intracellular source of contrast. We then demonstrated that those structures can indeed be tracked and quantified in a variety of unstained tissues using THG microscopy. Finally, we applied THG microscopy to the visualization of early development of live unstained Drosophila embryos. On this model, we studied mechanisms of induced phototoxicity and demonstrated that THG microscopy can provide a complete 3D description of morphogenetic movements in wild-type and mutant embryos without perturbation.
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Submitted on : Wednesday, July 28, 2010 - 3:41:40 PM
Last modification on : Thursday, June 4, 2020 - 2:10:04 PM
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  • HAL Id : pastel-00001978, version 1



Delphine Débarre. Microscopie par génération de troisième harmonique appliquée à la biologie.. Biophysique []. Ecole Polytechnique X, 2006. Français. ⟨pastel-00001978⟩



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