Analytical Chemistry Commons^™

Lipid Rafts, Exosomal Vesicles And Anti-Giardial Therapies, Brian Ivan Grajeda

Open Access Theses & Dissertations

Giardia lamblia, a protozoan parasite, is a major cause of waterborne infection, worldwide. While the trophozoite form of this parasite induces pathological symptoms in the gut, the cyst forms transmit the infection via contaminated water. Since Giardia is a non-invasive parasite, the actual mechanism by which it causes infection remains elusive. We have previously reported that Giardia assembles cholesterol and GM1 glycolipid-enriched lipid rafts (LRs) that participate in encystation and cyst production. To further delineate the role of LRs in pathogenesis, we isolated LRs from Giardia and subjected them to proteomic analysis. Various cellular proteins including the virulent proteins—e.g., giardins, …

Validation Of A Deployable Proteomic Assay For The Serological Screening Of Sexual Assault Samples, Catherine O'Sullivan Brown

Electronic Theses and Dissertations

Protein mass spectrometry (MS) has emerged as a technique to supplant traditional serological tests for body fluid identification. It was hypothesized that proteomic techniques would surpass the sensitivity and specificity of traditional serological techniques. An automated workflow coupled with protein MS has been developed for the confirmatory identification of five biological fluids. A developmental validation was completed, assessing parameters such as reproducibility, sensitivity, ion suppression, and limit of detection. Implementation was determined through tandem sample processing by MS, traditional serological tests, and standard DNA profiling methods. The MS approach offered superior detection limits while also providing true confirmatory results, producing …

Deep-Ultraviolet Laser Ablation For Biomolecular Analysis, Oluwaremilekun Omowunmi Lawal

LSU Doctoral Dissertations

New sampling and ionization techniques were developed using a deep-ultraviolet (DUV) laser at 193 nm wavelength for analysis of large biomolecules. A micrometer spatial resolution DUV laser ablation and capture system was constructed to sample tissue biomolecules. The spot size measurements of the focused beam showed that subcellular spots with ca. 2 μm diameter can be readily achieved with this wavelength. To demonstrate the extraction of tissue biomolecules with the DUV laser, regions of tissue sections were ablated and captured for offline proteomic and genomic analysis. The proteomic studies revealed that the tissue proteins were ejected without fragmentation and that …

Intrinsic Buffer Hydroxyl Radical Dosimetry For Hydroxyl Radical Protein Footprinting, Addison Roush

Honors Theses

Hydroxyl radical protein footprinting (HRPF) coupled to mass spectrometry is a powerful technique for the analysis of protein topography as it generates covalent mass labels that can survive downstream sample handling, and it is sensitive to the solvent accessibility of amino acid sidechains. Of the multiple platforms for HRPF, fast photochemical oxidation of proteins (FPOP) utilizes a pulsed 248 nm KrF excimer laser to label proteins by photolyzing hydrogen peroxide. FPOP is the most widely used HRPF platform because it labels proteins faster than unfolding can occur. Variations in FPOP sample conditions make it difficult to compare results between experiments …

An Exploration Of Protein And Dna Components In Fingerprint Residue, Ashley Borrego

Student Theses

The main focus of this project was to investigate the protein and DNA components in both sebaceous and eccrine fingerprints. This study investigated the relative content of DNA and proteins in eccrine fingerprints to sebaceous fingerprints. All volunteers were instructed to wash and dry their hands prior to depositing parallel thumbprints. Twenty volunteers were instructed to touch their face to produce sebaceous prints, and 5 volunteers were instructed to wear gloves over a heat source to produce sweaty or eccrine prints. Microscopy was used to score the cellular debris of the right fingerprint on a scale of 1-4 based on …

Enhanced Mass Spectrometric Analysis Of Peptides And Proteins Using Polymeric Reverse Micelles, Mahalia Adelina Corazon Paningbatan Serrano

Doctoral Dissertations

Mass spectrometry (MS) has become a key and indispensable tool in the identification, characterization, and quantitative analysis of proteins owing to its universality, sensitivity, specificity, and its capability for multiplexed detection. Because biological samples containing these protein analytes are almost always complex systems, various techniques are employed in conjunction with MS to fully harness its analytical potential and enhance its detection capabilities. This dissertation explores the use of amphiphilic polymeric reverse micelles in enriching proteins and peptides from complex biological mixtures and in enhancing their mass spectrometric analysis. Fundamental studies that elucidate the molecular basis for the observed MS signal …

Xflow: An Algorithm For Extracting Ion Chromatograms, Mathew M. Gutierrez, Rob Smith

Graduate Student Theses, Dissertations, & Professional Papers

Abstract: Mass spectrometry is a fundamental tool for modern proteomics. The increasing availability of mass spectrometry data paired with the increasing sensitivity and fidelity of the instruments necessitates new and more potent analytical methods. To that end, we have created and present XFlow, a feature detection algorithm for extracting ion chromatograms from MS1 LC-MS data. XFlow is a parameter-free procedurally agnostic feature detection algorithm that utilizes the latent properties of ion chromatograms to resolve them from the surrounding noise present in MS1 data. XFlow is designed to function on either profile or centroided data across different resolutions and instruments. This …

Analytical Method Development For The Detection And Analysis Of Protein Carbonyls, Chelsea M. Coffey

Theses and Dissertations

Oxidative stress can result in changes to many biomolecules and also affect their activities. We are interested in protein carbonylation, a type of unnatural oxidation which has been associated with numerous degenerative disease states and is also a consequence of the natural aging process. Protein carbonyls are stable species, but countless analytical barriers exist in terms of their identification. Thus, the main goal of this work was to develop and optimize analytical methods that could be used to help us better understand which, where, and how proteins are being carbonylated.

Initial studies involved method validation for carbonylating, tagging, and enriching …

Identification And The Significance Of Selective Proteins In Bile And Plasma Of Normal And Health-Compromised Chickens, Balamurugan Packialakshmi

Graduate Theses and Dissertations

During the last 50 years, animal breeding programs in commercial poultry have made significant progress in the bodyweight gain of broilers but led to several metabolic and skeletal disorders. Lameness associated with proximal femur known as femoral head separation (FHS) or femoral head necrosis (FHN) is one of the major metabolic disorders in poultry industry. In order to select for healthy chickens, markers that can distinguish between healthy and affected birds are required. Biomarkers from blood represent an ideal and rich source of markers which can be obtained using minimally invasive methods. The biomarkers were explored in an experimental model …

Demonstration Of A Targeted Proteome Characterization Approach For Examining Specific Metabolic Pathways In Complex Bacterial Systems, Adam Justin Martin

Masters Theses

Multiple Reaction Monitoring (MRM) is a powerful tandem mass spectrometry (MS/MS) tool frequently implemented in proteomic studies to provide targeted analysis of proteins and peptides. The selectivity that MRM delivers is so strong that it provides the quadrupole mass spectrometers (QQQ), on which it is commonly employed, with pertinence to proteomic studies that they would otherwise lack for their relatively low resolution. Additionally, this increased level of selectivity is sufficient enough to supplant complicated fractionation techniques, additional dimensions of chromatography, and 24 hour long MS/MS experiments in simplistic biological samples. But there is a deficiency of evidence to determine the …

Development Of A Novel Electron-Transfer Secondary Reaction Matrix, Characterization Of The Site–Specificity Of Novel Bilin-Lyase, And Fundulus Grandis Protein Expression Investigation Using Mass Spectrometry, Mohamed N. Boutaghou

University of New Orleans Theses and Dissertations

Reported in this dissertation are the results of investigations performed at the New Orleans Center for Mass Spectrometry at the University of New Orleans. The projects that are detailed in the coming pages take on a variety of subjects, but a common thread is that each employs matrix-assisted laser desorption/ionization (MALDI) mass spectrometry to solve a problem. Fundamental aspects of MALDI in-plume ionization are implicated in the introduction of a newly developed electron-transfer secondary ionization matrix. The remaining projects are related to the ever expanding field of proteomics. Mass spectrometry was used to investigate the site specificity of a newly …

Advanced Techniques In Mass Spectrometry For Qualitative And Quantitative Protein Characterization, Andrew Boissy Dykstra

Doctoral Dissertations

Though mass spectrometry has earned a central role in the field of proteomics due to its versatility in a wide range of experiments, challenges and complications are still encountered when using mass spectrometry to characterize protein structures, post-translational modifications (PTMs), and abundances. In this dissertation, analytical methods utilizing mass spectrometry have been developed to address challenges associated with both qualitative and quantitative protein characterization. The effectiveness of using multiple pepsin-like proteases, both separately and in mixtures, combined with online proteolysis using a special triaxial probe has been demonstrated on an amyloid beta peptide related to the onset of Alzheimer’s disease. …

Multidimensional Liquid Chromatography Separations, Jacob N. Fairchild

Doctoral Dissertations

Many mixtures important to research consist of hundreds or even thousands of individual components of interest. These types of mixtures are far too complex to separate by a single chromatographic dimension in any reasonable amount of time. However, if a multidimensional approach is used, where a complex mixture is separated by an initial dimension, simpler fractions of that separation are collected and each of those fractions are analyzed individually, highly complex mixtures can be resolved in relatively short amounts of time. This dissertation serves as a guide to multidimensional chromatography, in particular, two-dimension liquid chromatography. There are many aspects of …