SUN ETD - Theses and Dissertations

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Browsing SUN ETD - Theses and Dissertations by Author "Badrodien, Imraan"

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    Two-Photon Light-Sheet Fluorescence Microscopy with Compressed Supercontinuum Pulses
    (Stellenbosch University, 2024-12) Badrodien, Imraan; Bosman, G. W.; Neethling, P. H.; Stellenbosch University. Faculty of Science. Dept. of Physics.
    Biological imaging has been transformed by the use of light-sheet fluorescence microscopy, which enables faster three-dimensional imaging with lower photobleaching and photo-toxicity compared to traditional microscopy. Nonlinear optical processes, specifically two-photon excitation fluorescence, have further improved imaging capabilities by enabling deeper penetration depths and reduced out-of-focus fluorescence. However, t he h igh p eak i ntensities required for nonlinear responses can lead to photodamage, making it imperative to maintain incident radiation below the damage threshold of biological samples when implementing nonlinear microscopy techniques. Recent advancements in all-normal-dispersion photonic crystal fibres (ANDi-PCF) have enabled the generation of highly stable broadband supercontinuum pulses that can be temporally compressed. While the multiphoton intrapulse interference phase scanning technique has been the popular choice for pulse characterisation, an alternative technique called i2PIE has been proposed and developed to reconstruct the spectral phase of a pulse. This dissertation reports the use of broadband supercontinuum pulses generated in an ANDi-PCF, compressed using the i2PIE phase measurement technique, in two-photon light-sheet microscopy. The performance is compared with other pulse compression techniques, and the application of this new method in imaging biological samples is demonstrated. The results indicate that the i2PIE technique enhanced the imaging capabilities of the two-photon light-sheet microscope, producing increased contrast and signal-to-noise ratios across all images compared to traditional compression techniques such as MIIPS or compression employing only the use of chirped mirrors. This demonstrates the potential of the i2PIE spectral phase measurement technique to enhance the imaging capabilities of two photon light-sheet fluorescence microscopy.