The Development and Characterization of Biologically-Inert Raman Reporter(s) for Live-cell Probing of Protein Trafficking and Localization Mediated by Posttranslational Modifications

Janelle Weslyn Salameh (Tami Kinzer-Ursem, advisor)

Cellular processes are maintained and regulated by complex protein networks through interactions with signaling molecules, metabolic modifications with unique functional groups, or direct protein-protein interactions. In the past decade, we gained tremendous insight into cellular events through the development of fluorescence labeling techniques such as genetically-encoded fluorescent proteins, immunostaining, and molecular dyes. However, these labeling techniques suffer from major drawbacks such as nonspecific interactions, cytotoxicity, and interference with native cellular processes. Additionally, conventional imaging techniques require invasive staining, contrast-enhancing procedures that only provide snap-shots of cellular events.

Posttranslational modification (PTM) of proteins is one of the most versatile approaches for regulating cellular functions. One of these modifications is protein lipidation in which proteins are covalently tagged with a variety of lipids. In recent years, these native protein modifications were widely exploited yielding a new generation of labeling technology, known as Click-iT labeling. This new technology allows for site-specific labeling of proteins, thus providing high sensitivity and low background signal by eliminating nonspecific interactions. However, Click-iT labeling relies on the introduction of fluorophores to the system, which perturbs the native environment of the cell and requires cell fixation prior to imaging. My research bypasses the need for fluorophores by utilizing existent protein tags and simply modifying these tags with small functional groups that exhibit distinct “fingerprints” when probed spectroscopically. I will be discussing the development a fatty acid analog with distinct Raman-active functional groups, and how this tag can be used to study protein trafficking and localization with little to no sample preparation steps.

A Fiber-Delivered Optoacoustic Guide for Precision Breast-Conserving Surgery

Lu Lan (Ji-Xin Cheng, advisor)

Breast-conserving surgery is a well-accepted breast cancer treatment, in which a tumor or a lump is removed along with sufficient surrounding normal tissue. A critical factor resulting in the current high re-operation rate is a lack of accurate real-time surgical guidance to locate the exact cancerous area and confirm excision of sufficient margins. In order to improve the efficiency and accuracy of BCS and to help reduce re-operation rate, we demonstrate a fiber-delivered optoacoustic guide, which mimics the traditional wire localization while providing additional real-time quantitative information about tumor location and margin. A nano-composite sphere of zinc-oxide nanoparticles and epoxy is formed on the proximal end of a multimode optical fiber to diffuse the light. The composite is then coated by a graphite layer to convert the light into an omnidirectional optoacoustic source when excited by a nanosecond pulsed laser. The optoacoustic signal generated has a high dynamic range (~55 dB) and spreads in a large apex angle of 260 degrees. By integrating an ultrasound detector into a surgical blade and using real-time visual guidance software, we demonstrate that this optoacoustic guide fiber could provide real-time tumor removal guidance. Furthermore, we present visualization of the tip of the guide wire using an augmented reality method to facilitate the intraoperative surgical planning, potentially improving the excision accuracy and efficiency.

***Bring your lunch to seminar – BMEGSA will provide snacks and drinks***