Optics Commons^™

Fundamental Limits Of Measuring Single-Molecule Rotational Mobility, Oumeng Zhang, Matthew D. Lew

Electrical & Systems Engineering Publications and Presentations

Various methods exist for measuring molecular orientation, thereby providing insight into biochemical activities at nanoscale. Since fluorescence intensity and not electric field is detected, these methods are limited to measuring even-order moments of molecular orientation. However, any measurement noise, for example photon shot noise, will result in nonzero measurements of any of these even-order moments, thereby causing rotationally-free molecules to appear to be partially constrained. Here, we build a model to quantify measurement errors in rotational mobility. Our theoretical framework enables scientists to choose the optimal single-molecule orientation measurement technique for any desired measurement accuracy and photon budget.

In Vivo Vascular Imaging With Photoacoustic Microscopy, Hsun-Chia Hsu

McKelvey School of Engineering Graduate Student Theses & Dissertations

Photoacoustic (PA) tomography (PAT) has received extensive attention in the last decade for its capability to provide label-free structural and functional imaging in biological tissue with highly scalable spatial resolution and penetration depth. Compared to modern optical modalities, PAT offers speckle-free images and is more sensitive to optical absorption contrast (with 100% relative sensitivity). By implementing different regimes of optical wavelength, PAT can be used to image diverse light-absorbing biomolecules. For example, hemoglobin is of particular interest in the visible wavelength regime owing to its dominant absorption, and lipids and water are more commonly studied in the near-infrared regime.

In …

Super‐Resolution Imaging Of Amyloid Structures Over Extended Times By Using Transient Binding Of Single Thioflavin T Molecules, Kevin Spehar, Tianben Ding, Yuanzi Sun, Niraja Kedia, Jin Lu, George R. Nahass, Matthew D. Lew, Jan Bieschke

Electrical & Systems Engineering Publications and Presentations

Oligomeric amyloid structures are crucial therapeutic targets in Alzheimer's and other amyloid diseases. However, these oligomers are too small to be resolved by standard light microscopy. We have developed a simple and versatile tool to image amyloid structures by using thioflavin T without the need for covalent labeling or immunostaining. The dynamic binding of single dye molecules generates photon bursts that are used for fluorophore localization on a nanometer scale. Thus, photobleaching cannot degrade image quality, allowing for extended observation times. Super‐resolution transient amyloid binding microscopy promises to directly image native amyloid by using standard probes and record amyloid dynamics …

Imaging The Three-Dimensional Orientation And Rotational Mobility Of Fluorescent Emitters Using The Tri-Spot Point Spread Function, Oumeng Zhang, Jin Lu, Tianben Ding, Matthew D. Lew

Electrical & Systems Engineering Publications and Presentations

Fluorescence photons emitted by single molecules contain rich information regarding their rotational motions, but adapting single-molecule localization microscopy (SMLM) to measure their orientations and rotational mobilities with high precision remains a challenge. Inspired by dipole radiation patterns, we design and implement a Tri-spot point spread function (PSF) that simultaneously measures the three-dimensional orientation and the rotational mobility of dipole-like emitters across a large field of view. We show that the orientation measurements done using the Tri-spot PSF are sufficiently accurate to correct the anisotropy-based localization bias, from 30 nm to 7 nm, in SMLM. We further characterize the emission anisotropy …

Fluorescence Guided Tumor Imaging: Foundations For Translational Applications, Jessica P. Miller

McKelvey School of Engineering Graduate Student Theses & Dissertations

Optical imaging for medical applications is a growing field, and it has the potential to improve medical outcomes through its increased sensitivity and specificity, lower cost, and small instrumentation footprint as compared to other imaging modalities. The method holds great promise, ranging from direct clinical use as a diagnostic or therapeutic tool, to pre-clinical applications for increased understanding of pathology. Additionally, optical imaging uses non-ionizing radiation which is safe for patients, so it can be used for repeated imaging procedures to monitor therapy, guide treatment, and provide real-time feedback. The versatile features of fluorescence-based optical imaging make it suited for …

Novel Sensing Mechanisms For Chemical And Bio-Sensing Using Whispering Gallery Mode Microresonators, He Huang

McKelvey School of Engineering Graduate Student Theses & Dissertations

Due to their ultra-high quality factor and small mode volume, whispering gallery mode (WGM) microresonators have proven to have exceptional sensing capabilities, with single particle level sensitivity to virions, proteins, and nucleic acids. Current sensing mechanisms rely on measuring the changes in the transmission spectrum of the resonator upon adsorption of the analyte on the surface of the resonator, appearing as either shift, splitting, or broadening of the resonance mode, all of which measure the polarizability of adsorbed analytes. In this dissertation, we present two new sensing mechanisms for WGM microresonators: the measurement of a dynamic chemical reaction around the …

Developing Photoacoustic Tomography Devices For Translational Medicine And Basic Science Research, Tsz Wai Wong

McKelvey School of Engineering Graduate Student Theses & Dissertations

Photoacoustic (PA) tomography (PAT) provides volumetric images of biological tissue with scalable spatial resolutions and imaging depths, while preserving the same imaging contrast—optical absorption. Taking the advantage of its 100% sensitivity to optical absorption, PAT has been widely applied in structural, functional, and molecular imaging, with both endogenous and exogenous contrasts, at superior depths than pure optical methods. Intuitively, hemoglobin has been the most commonly studied biomolecule in PAT due to its strong absorption in the visible wavelength regime.

One of the main focuses of this dissertation is to investigate an underexplored wavelength regime—ultraviolet (UV), which allows us to image …

Developing Wavefront Shaping Techniques For Focusing Through Highly Dynamic Scattering Media, Ashton Hemphill

McKelvey School of Engineering Graduate Student Theses & Dissertations

One of the prime limiting factors of optical imaging in biological applications is the diffusion of light by tissue, which prevents focusing at depths greater than the optical diffusion limit of ~1 mm in soft tissue. This greatly restricts the utility of optical diagnostic and therapeutic techniques, such as optogenetics, microsurgery, optical tweezing, and phototherapy of deep tissue, which require focused light in order to function. Wavefront shaping extends the depth at which optical focusing may be achieved by compensating for phase distortions induced by scattering, allowing for focusing through constructive interference.

However, due to physiological motion, scattering of light …

Measuring 3d Molecular Orientation And Rotational Mobility Using A Tri-Spot Point Spread Function, Oumeng Zhang, Tianben Ding, Jin Lu, Hesam Mazidi, Matthew D. Lew

Electrical & Systems Engineering Publications and Presentations

We present a method to measure the molecular orientation and rotational mobility of single-molecule emitters by designing and implementing a Tri-spot point spread function. It can measure all degrees of freedom related to molecular orientation and rotational mobility. Its design is optimized by maximizing the theoretical limit of its measurement precision. We evaluate the precision and accuracy of the Tri-spot PSF by measuring the orientation and effective rotational mobility of single fluorescent molecules embedded in a polymer matrix.

Multiscale Imaging Of The Mouse Cortex Using Two-Photon Microscopy And Wide-Field Illumination, Jonathan Richard Bumstead

McKelvey School of Engineering Graduate Student Theses & Dissertations

The mouse brain can be studied over vast spatial scales ranging from microscopic imaging of single neurons to macroscopic measurements of hemodynamics acquired over the majority of the mouse cortex. However, most neuroimaging modalities are limited by a fundamental trade-off between the spatial resolution and the field-of-view (FOV) over which the brain can be imaged, making it difficult to fully understand the functional and structural architecture of the healthy mouse brain and its disruption in disease. My dissertation has focused on developing multiscale optical systems capable of imaging the mouse brain at both microscopic and mesoscopic spatial scales, specifically addressing …

Measuring Molecular Orientation And Rotational Mobility Using A Tri-Spot Point Spread Function, Oumeng Zhang

McKelvey School of Engineering Graduate Student Theses & Dissertations

Single molecules have become a powerful tool for biophysicists since they were first optically detected 28 years ago. Understanding molecular orientation can not only improve the accuracy of single-molecule localization, but it can also provide insight into biochemical behaviors at the nanoscale. In this thesis, I present a method to measure the molecular orientation and rotational mobility of single-molecule emitters by designing and implementing a tri-spot point spread function. The point spread function is designed so that it is capable of measuring all degrees of freedom related to molecular orientation and rotational mobility. Its design is optimized by maximizing the …

Goggle Augmented Imaging And Navigation System For Fluorescence-Guided Surgery, Suman Bikash Mondal

McKelvey School of Engineering Graduate Student Theses & Dissertations

Surgery remains the only curative option for most solid tumors. The standard-of-care usually involves tumor resection and sentinel lymph node biopsy for cancer staging. Surgeons rely on their vision and touch to distinguish healthy from cancer tissue during surgery, often leading to incomplete tumor resection that necessitates repeat surgery. Sentinel lymph node biopsy by conventional radioactive tracking exposes patients and caregivers to ionizing radiation, while blue dye tracking stains the tissue highlighting only superficial lymph nodes. Improper identification of sentinel lymph nodes may misdiagnose the stage of the cancer. Therefore there is a clinical need for accurate intraoperative tumor and …

Optical Resonators And Fiber Tapers As Transducers For Detection Of Nanoparticles And Bio-Molecules, Huzeyfe Yilmaz

McKelvey School of Engineering Graduate Student Theses & Dissertations

In recent years, detection of biological interactions on single molecule level has aspired many researchers to investigate several optical, chemical, electrical and mechanical sensing tools. Among these tools, toroidal optical resonators lead the way in detection of the smallest particle/molecule with the real time measurements. In this work, bio-sensing capabilities of toroidal optical resonators are investigated. Bio-sensing is realized via measuring the analyte-antigen interaction while the antigen is immobilized through a novel functionalization method.

Not long ago, detection of single nanoparticles using optical resonators has been accomplished however the need for cost-effective and practical transducers demands simpler tools. A tapered …