Structured illumination for in-vivo sectioning and imaging

Abstract

Structured illumination microscopy (SIM) is a method of non-invasively extracting depth information about a specimen. By using modulated light focused at a discrete depth, it is able optically section a single plane of focus, even in the presence of scattered light from other, out-of-focus planes. However, in order for images to be accurately extracted, the modulation pattern must be positioned into three discrete phases, whose alignment is critical to artifact-free reconstruction. Due to this constraint, SIM does not work well at depth, within optically turbid media, such as human skin. Our research considers two new methods of applying and processing SIM. Random modulation patterns are considered which do not rely on discrete phase differences and are therefore much more robust at depth. In parallel, a complimentary super-resolution method is applied to extract both depth information, as well as enhanced resolution that exceeds the diffraction limit of the system. Secondly, a new single image processing scheme, based on the Hilbert transform is developed to process traditional discrete frequency, modulation patterns. This method mitigates the need for phase alignment greatly increasing the depth of the sectioning, as well as allowing for real-time processing. Further, we produce these images using an LED source in reflectance, obviating the need for fluorescent markers, which we are able to demonstrate by producing in-vivo images on a human subject. This research extends the depth of SIM to ∼100um within a tissue sample bringing it much closer to other clinical tools, such as Confocal, at a fraction of the cost. i 11 Dissertation Conclusion 122 v List of Figures Diagram outlining a typical Confocal reflectance system. This only shows the pin-hole system, which needs to be scanned in the X and Y directions to build a full two dimensional image.. .. 2 Diagram demonstrating the high frequency modulation pattern at the focal plane. Regions above and below the focal plane quickly go out of focus, removing the modulation pattern.. .. .. . .