Application of optical coherence tomography in investigating cell migration

Chemotaxis and cell migration are important processes for life, involved in organism development and homeostasis and implicated in a number of disease states. Dictyostelium discoideum, an amoeba, is a useful model for investigation of chemotaxis and development, due to its ability to undergo chemotactic aggregation and development upon starvation. Although cell migration has been well described on planar transparent surfaces, it is uncertain how well these conditions replicate the natural environment of a cell. However, attempts to better replicate these environments generally make use of opaque substrates and 3D matrices, in which it is more challenging to image cell migration. Protocols were developed to enable optical coherence tomography, a 3D structural imaging technique which requires no sample processing or staining, to be successfully employed in imaging Dictyostelium cell migration in time-lapse on non-transparent substrata and within an agarose gel. I compared the effects of two substrates, a nitrocellulose filter and a polystyrene Petri dish on aggregating cells and found differences in speed but not persistence. Extension of this to include cells within agarose revealed that these cells exhibited less directed migration, but their velocity was unaffected. I showed that cells lacking myosin II failed to complete development within an agarose gel and had significantly reduced velocity and directional migration when compared to their parent strain. Furthermore, the velocities of cells migrating within agarose gel were bimodally distributed, potentially indicating two distinct cell populations, fast and slow, and fast movement was shown to be largely myosin II dependent. Great potential therefore exists for cell-substrate and cell-matrix interactions to affect the migration character of cells, even those, such as Dictyostelium, which do not form strong focal adhesions. Moreover a properly ordered cytoskeleton is implicated in enabling cells to effectively utilise different modes of cell motility

Quantum Coherent Dynamics in a dc SQUID Phase Qubit Using an LC Filter

A dc SQUID phase qubit consists of two Josephson junctions in a loop. One junction acts as a qubit with two lowest energy levels forming the |0> and |1> status. The second junction and the loop inductance act to isolate the qubit junction from noise. In this thesis, I report on the improvement of the relaxation time and the coherence time in a dc SQUID phase qubit that used an LC filter. I also report the measurement of anomalous switching curves. In order to improve the relaxation and coherence times, I used two isolation networks, an LC isolation network and an inductive isolation network, to decouple the device from the current bias lines. This produced a very large total effective resistance of the input leads that increases the relaxation time of the qubit. In addition, I connected a low-loss SiNx shunting capacitor across the qubit junction to reduce dielectric losses. I measured two dc SQUID phase qubits. Device DS6 had a 4 (μm)2 Al/AlOx/Al qubit junction with a critical current of 0.5 μA and a 1 pF shunting capacitor. It used an LC filter made from a 10 nH inductor and a 145 pF capacitor. The capacitors contained N-H rich SiNx which produced a loss tangent of about 7×10-4. Device DS8 had a 2 (μm)2 Al/AlOx/Al qubit junction with a critical current of 77 nA and an LC filter similar to the first one. The shunting capacitor contained Si-H rich SiNx. Using a pulse readout technique, I measured the characteristics of the qubits, including the transition spectrum, Rabi oscillations, relaxation, Ramsey fringes and state tomography. The best relaxation time T1 for device DS6 was 32 ns and 280 ns for device DS8. The best Rabi decay time T' for DS6 was 42 ns while for device DS8 it was 120 ns. From these and other data I obtained estimates for the best coherence time T2 in device DS6 of 61 ns and 76 ns in device DS8. In DS8, I observed anomalous switching curves; i.e. switching curves which were qualitatively different from conventional switching curves. In the conventional case, the switching curve for the superposition state is the weighted sum of the |0> and |1> curves, but it was not in device DS8. Instead, the switching curve shifted along the current axis as the exited state probability increased. I present a model for understanding the behavior and use this model to extract the probability to be in the excited state

Hard X-ray grazing incidence ptychography: Large field-of-view nanostructure imaging with ultra-high surface sensitivity

We demonstrate a technique that allows highly surface sensitive imaging of nanostructures on planar surfaces over large areas, providing a new avenue for research in materials science, especially for \textit{in situ} applications. The capabilities of hard X-ray grazing incidence ptychography combine aspects from imaging, reflectometry and grazing incidence small angle scattering in providing large field-of-view images with high resolution transverse to the beam, horizontally and along the surface normal. Thus, it yields data with resolutions approaching electron microscopy, in two dimensions, but over much larger areas and with a poorer resolution in the third spatial dimension, along the beam propagation direction. Similar to grazing incidence small angle X-ray scattering, this technique facilitates the characterization of nanostructures across statistically significant surface areas or volumes within potentially feasible time frames for \textit{in situ} experiments, while also providing spatial information.Comment: 8 pages, 6 figure

Removing striping artifacts in light-sheet fluorescence microscopy: a review

In recent years, light-sheet fluorescence microscopy (LSFM) has found a broad application for imaging of diverse biological samples, ranging from sub-cellular structures to whole animals, both in-vivo and ex-vivo, owing to its many advantages relative to point-scanning methods. By providing the selective illumination of sample single planes, LSFM achieves an intrinsic optical sectioning and direct 2D image acquisition, with low out-of-focus fluorescence background, sample photo-damage and photo-bleaching. On the other hand, such an illumination scheme is prone to light absorption or scattering effects, which lead to uneven illumination and striping artifacts in the images, oriented along the light sheet propagation direction. Several methods have been developed to address this issue, ranging from fully optical solutions to entirely digital post-processing approaches. In this work, we present them, outlining their advantages, performance and limitations

Removing striping artifacts in light-sheet fluorescence microscopy: a review

In recent years, light-sheet fluorescence microscopy (LSFM) has found a broad application for imaging of diverse biological samples, ranging from sub-cellular structures to whole animals, both in-vivo and ex-vivo, owing to its many advantages relative to point-scanning methods. By providing the selective illumination of sample single planes, LSFM achieves an intrinsic optical sectioning and direct 2D image acquisition, with low out-of-focus fluorescence background, sample photo-damage and photo-bleaching. On the other hand, such an illumination scheme is prone to light absorption or scattering effects, which lead to uneven illumination and striping artifacts in the images, oriented along the light sheet propagation direction. Several methods have been developed to address this issue, ranging from fully optical solutions to entirely digital post-processing approaches. In this work, we present them, outlining their advantages, performance and limitations

“Applications of Coherent Electron Beams

The use of coherent beams for interferometric measurements has gained great popularity in light optics over the last several decades. The availability of coherent electron sources has now opened the door to apply the concept of holographic imaging in many new areas. Off-axis holograms can now be recorded in field emission transmission electron microscopes equipped with the electron optical equivalent of a biprism. This technique allows the accurate retrieval of phase and amplitude of the electron wave, which has been transmitted through a sample. The sensitivity of the phase of the electron wave to electrical potentials makes it possible to map out potential distributions on the specimen with sub-micron resolution. As part of this thesis, off-axis electron holography has been applied to map out the small potential changes, which occur over pn-junctions in doped semiconductor devices. To this end a special alignment of the electron microscope has been devised, and new methods for preparing electron transparent samples of semiconductor devices, specially tailored for electron holography, have been developed. Also, new ways to improve on the existing electron biprism technology have been investigated. The observed voltage signals could be linked to active dopants in silicon by annealing experiments, and the viability of the method for voltage profiling of real-world semiconductor devices has been demonstrated. The use of coherent electron emitters also allows the recording of in-line electron holograms in a lens-less projection microscope at ultra-low beam energies. Such a point projection microscope, which is capable of recording in-line holograms in transmission imaging, has been built. With defect review on silicon wafers as a possible application for in-line electron holography in mind, the feasibility of point projection imaging in a reflection geometry has been demonstrated. In this context the elastic backscattered yield for electrons in different materials and under different geometries has been calculated using Monte Carlo simulations. Several problems, which occur in reflection imaging, are pointed out and possible solutions are presented

Functional photoacoustic tomography of animal brains

This research is primarily focused on laser-based non-invasive photoacoustic tomography of small animal brains. Photoacoustic tomography, a novel imaging modality, was applied to visualize the distribution of optical absorptions in small-animal brains through the skin and skull. This technique combines the high-contrast advantage of optical imaging with the high-resolution advantage of ultrasonic imaging. Based on the intrinsic optical contrast, this imaging system successfully visualized three-dimensional tissue structures in intact brains, including lesions and tumors in brain cerebral cortex. Physiological changes and functional activities in brains, including cerebral blood volume and blood oxygenation in addition to anatomical information, were also satisfactorily monitored. This technique successfully imaged the dynamic distributions of exogenous contrast agents in small-animal brains. Photoacoustic angiography in small-animal brains yielding high contrast and high spatial resolution was implemented noninvasively using intravenously injected absorbing dyes. In the appendix, the theory of Monte Carlo simulation of polarized light propagation in scattering media was briefly summarized

Information multiplexing from optical holography to multi-channel metaholography

Holography offers a vital platform for optical information storage and processing, which has a profound impact on many photonic applications, including 3D displays, LiDAR, optical encryption, and artificial intelligence. In this review, we provide a comprehensive overview of optical holography, moving from volume holography based on optically thick holograms to digital holography using ultrathin metasurface holograms in nanophotonics. We review the use of volume holograms for holographic multiplexing through the linear momentum selectivity and other approaches and highlight the emerging use of digital holograms that can be implemented by ultrathin metasurfaces. We will summarize the fabrication of different holographic recording media and digital holograms based on recent advances in flat meta-optics and nanotechnology. We highlight the rapidly developing field of metasurface holography, presenting the use of multi-functional metasurfaces for multiplexing holography in the use of polarization, wavelength, and incident angle of light. In the scope of holographic applications, we will focus on high bandwidth metasurface holograms that offer the strong sensitivity to the orbital angular momentum of light. At the end, we will provide a short summary of this review article and our perspectives on the future development of the vivid holography field.AM and AB acknowledge support from projects PROMETEO/2021/006 and IDIFEDER/2021/014 (Generalitat Valenciana, Spain; cofunded by European Union through the FEDER Programme) and PID2021-123124OB-I00 (Ministerio de Ciencia e Innovación, Spain). This work was funded by the Australian Research Council (DE220101085, DP220102152). S.A.M. further acknowledges the EPSRC (EP/W017075/1) and the Lee-Lucas Chair in Physics