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Meyer, Verena. "Selective analysis of specific HLA ligand repertoires: poxviral CD8+ T cell epitopes and phosphorylated HLA ligands of tumor cells." [S.l. : s.n.], 2008.
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Polyak, Maria J. "Structural analysis and epitope mapping of CD20." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape4/PQDD_0020/MQ49652.pdf.
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Paul, Sinu. "Host-pathogen interactions and evolution of epitopes in HIV-1: understanding selection and escape." Kent State University / OhioLINK, 2012. http://rave.ohiolink.edu/etdc/view?acc_num=kent1334509644.
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Bothamley, Graham Henry. "Analysis of epitope-specific antibody levels in tuberculosis." Thesis, Imperial College London, 1990. http://hdl.handle.net/10044/1/47780.
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Behr, Jonathan Robert. "Novel tools for sequence and epitope analysis of glycosaminoglycans." Thesis, Massachusetts Institute of Technology, 2007. http://hdl.handle.net/1721.1/42383.
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Thesis (Ph. D.)--Massachusetts Institute of Technology, Biological Engineering Division, 2007.<br>Includes bibliographical references.<br>Our understanding of glycosaminoglycan (GAG) biology has been limited by a lack of sensitive and efficient analytical tools designed to deal with these complex molecules. GAGs are heterogeneous and often sulfated linear polys accharides found throughout the extracellular environment, and available to researchers only in limited mixtures. A series of sensitive label-free analytical tools were developed to provide sequence information and to quantify whole epitopes from GAG mixtures. Three complementary sets of tools were developed to provide GAG sequence information. Two novel exolytic sulfatases from Flavobacterium heparinum that degrade heparan/heparan sulfate glycosaminoglycans (HSGAGs) were cloned and characterized. These exolytic enzymes enabled the exo-sequencing of a HSGAG oligosaccharide. Phenylboronic acids (PBAs) were specifically reacted with unsulfated chondroitin sulfate (CS) disaccharides from within a larger mixture. The resulting cyclic esters were easily detected in mass spectrometry (MS) using the distinct isotopic abundance of boron. Electrospray ionization tandem mass spectrometry (ESI-MSn) was employed to determine the fragmentation patterns of HSGAG disaccharides. These patterns were used to quantify relative amounts of isomeric disaccharides in a mixture. Fragmentation information is valuable for building methods for oligosaccharide sequencing, and the general method can be applied to quantify any isomers using MSn. Three other tools were developed to quantify GAG epitopes. Two microfluidic devices were characterized as HSGAG sensors. Sensors were functionalized either with protamine to quantify total HSGAGs or with antithrombin-III (AT-III) to quantify a specific anticoagulant epitope.<br>(cont.) A charge sensitive silicon field effect sensor accurately quantified clinically relevant anticoagulants including low molecular weight heparins (LMWH), even out of serum. A mass sensitive suspended microchannel resonator (SMR) measured the same clinically relevant HSGAGs. When these two sensors were compared, the SMR proved more robust and versatile. The SMR signal is more stable, it can be reused ad infinitum, and surface modifications can be automated and monitored. The field effect sensor provided an advantage in selectivity by preferentially detecting highly charged HSGAGs instead of any massive, non-specifically bound proteins. Lastly, anti-HSGAG single chain variable fragments (scFv) were evolved using yeast surface display towards generating antibodies for HSGAG epitope sensing and clinical GAG neutralization.<br>by Jonathan Robert Behr.<br>Ph.D.
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Patel, Sarju Dilipkumar. "Analysis of myelin-reactive T lymphocyte function in models of multiple sclerosis." Thesis, University of Edinburgh, 2008. http://hdl.handle.net/1842/3489.
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Immune tolerance to self antigens prevents the onset of autoimmune diseases such as Multiple Sclerosis (MS). There are three branches of tolerance which allow the auto-aggressive potential of T lymphocytes to be limited; these are death, anergy-adaptation and regulation. The main body of this work attempts to clarify a role for adaptation in maintaining the sensitivity of the autoreactive T cell repertoire below a ‘threshold for harm’ in the mouse model of MS, experimental autoimmune encephalomyelitis (EAE). The well defined myelin basic protein (MBP) Ac1-9 epitope altered peptide ligand (APL) system has been used to develop a model allowing the examination of mechanisms underlying the adaptation of cells. Previous data showed immunisation with the 4Lys (wild-type) epitope mediated disease whereas a superagonist APL with a tyrosine substitution at position 4 (4Tyr) did not, despite showing potency in vitro. This was shown to be a result of both activation induced cell death and adaptation. Here an in vitro model was developed using MBP-reactive TCR transgenic cells to make predictions about the mechanisms underlying adaptation. These data lead to the conclusion that T cells can adapt (become less sensitive) either before or after encounter with the wild-type peptide, leading to a reversal of their pathogenic potential. The MBP APL system and MBP reactive transgenic cells were also used to assess the contribution of epitope spreading in a relapsing-remitting (RR) model of EAE induced with proteolipid protein. The cells were tracked and changes in phenotype and behaviour were monitored. The data show that disease induced with one antigen can be manipulated with cells relevant to a different antigen and that bystander suppression may be an effective weapon in controlling the progression to RR-EAE.
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Ernstoff, Elana Ann. "HIV subtype C diversity: analysis of the relationship of sequence diversity to proposed epitope locations." Thesis, University of the Western Cape, 2002. http://etd.uwc.ac.za/index.php?module=etd&action=viewtitle&id=gen8Srv25Nme4_9247_1185441328.
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<p>Southern Africa is facing one of the most serious HIV epidemics. This project contributes to the HIVNET, Network for Prevention Trials cohort for vaccine development. HIVÂs biology and rapid mutation rate have made vaccine design difficult. We examined HIV-1 subtype C diversity and how it relates to CTL epitope location along viral gag sequences. We found a negative correlation between codon sites under positive selection and epitope regions<br>suggesting epitope regions are evolutionarily conserved. It is possible that epitopes exist in non-conserved regions, yet fail to be detected due to the reference strain diverging from the circulating viral population. To test if CTL clustering is an artifact of the reference strain, we calculated differences between the gag codons and the reference strain. We found a weak negative correlation, suggesting epitopes in less conserved regions maybe evading detection. Locating conserved and optimal epitopes that can be recognized by CTLs is essential for the design of vaccine reagents.</p>
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Costello, Ryan. "Towards the analyses of cell lineages using conditional gene alterations." Thesis, University of Manchester, 2016. https://www.research.manchester.ac.uk/portal/en/theses/towards-the-analyses-of-cell-lineages-using-conditional-gene-alterations(af6e20c3-1dd5-4c46-b14e-e062900b812a).html.
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The ability to precisely modify the mouse genome is an invaluable tool for any researcher. If an artificial epitope sequence is integrated into target loci in specific cell types, it is possible to generate mice with these cells specifically tagged with the epitope, which can be used for many subsequent studies. Homologous recombination and the Cre/loxP system have been used to generate targeted and conditional transgenic mice, which have provided the basis for many studies into gene function. However, in recent years, improvements in technology have led to the development of RNA and protein based methods of specifically editing DNA sequences at user-selected loci. This thesis aimed to investigate the effectiveness of the novel gene targeting methods TALEN and CRISPR/Cas9. It also aimed to utilize different strains of mice generated using the Cre/loxP system in Trichuris muris, an animal infection model of the human disease Trichuris trichiura. TALENs use a pair of protein-based monomers specific to the sense and anti-sense strand of a target DNA sequence to dimerise a FokI nuclease and initiate a deletion in the genome. As a study into the practical use of this emerging technology TALENs were generated to target Oct-4 (a stem cell marker) in order to integrate an artificial epitope sequence, which could be used for enrichment experiments. The CRISPR/Cas9 is a very efficient RNA-based system used for modifying a target sequence. This system has been utilized to integrate an epitope sequence into the Rosa26 locus, downstream of a floxed STOP codon. This allows for expression of the epitope following the introduction of tissue restricted Cre recombinase. IL-1 is an important cytokine in the immune response towards T.muris. IL-1R1 was conditionally removed in CD4 cells and the role of IL-1 signaling in developing Th1, Th2 and Th17 responses was then studied. Interestingly, IL-1R1fl/fl CD4Cre mice could generate Th1 and Th2 response but showed a reduction in IL-22 and IL-17 production, two key Th17 cytokines. Infected IL-1R1fl/fl CD4Cre mice also displayed increased gut morphology and goblet cell hyperplasia. Therefore, it was concluded that IL-1 signaling from CD4 cells has an important role in host defense and the development of a full Th17 response. It was also shown that removing IL-1R1 in naïve mice had no affect on lymphocyte development. IL-10 is an anti-inflammatory cytokine expressed by gut macrophages, which contributes to homeostatic control of the immune system. IL-10R was specifically removed in the macrophage specific Cre lines LysMCre and also in CX3CR1Cre as a way of comparing the two Cre drivers. The mice were then infected with T.muris and displayed significant inflammation and also failure to expel the worms in the LysMCre model. This suggests a role for this model in future studies of gut macrophages. Clearly, animal models are very important in the study of gene function and also as a method of assessing the application of new technologies such as CRISPR/Cas9. Future work with the artificial epitope specifically targeted into important cell lines will form the basis of many important studies directly applicable to human disorders. As the technologies improve, the scope for developing therapeutics increases and genetic modification has an immeasurable role to play.
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Dorea, Regina Coeli Cunha. "Immunity to murine cutaneous leishmaniasis : T-cell epitope analysis by Leishmania Mexicana-specific T-cell lines." Thesis, University of Strathclyde, 1990. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.291990.
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Mickael, Claudia Silva. "Real-time RT-PCR analysis of two epitope regions encoded by the VP2 gene of infectious bursal disease viruses." Connect to this title online, 2005. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1116532082.
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Thesis (Ph. D.)--Ohio State University, 2005.<br>Title from first page of PDF file. Document formatted into pages; contains xiii, 136 p.; also includes graphics (some col.) Includes bibliographical references (p. 120-136). Available online via OhioLINK's ETD Center
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Husslik, Felix [Verfasser], Harald [Akademischer Betreuer] Kolmar, and Vieths [Akademischer Betreuer] Stefan. "Analysis of the IgE epitope profile of the soybean allergen Gly m 4 / Felix Husslik ; Harald Kolmar, Vieths Stefan." Darmstadt : Universitäts- und Landesbibliothek Darmstadt, 2016. http://d-nb.info/1120014565/34.
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Rump, Kirsten [Verfasser], and Thomas [Akademischer Betreuer] Lengauer. "Computational immunology : analyses of viral escape, epitope binding and T cell receptor recognition / Kirsten Rump. Betreuer: Thomas Lengauer." Saarbrücken : Saarländische Universitäts- und Landesbibliothek, 2011. http://d-nb.info/1051326125/34.
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Rump, Kirsten Verfasser], and Thomas [Akademischer Betreuer] [Lengauer. "Computational immunology : analyses of viral escape, epitope binding and T cell receptor recognition / Kirsten Rump. Betreuer: Thomas Lengauer." Saarbrücken : Saarländische Universitäts- und Landesbibliothek, 2011. http://nbn-resolving.de/urn:nbn:de:bsz:291-scidok-42334.
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Lounsbach, Gillian Ruth. "Expression and epitopic analysis of the respiratory syncytial virus fusion protein in Escherichia coli." Thesis, University of Newcastle Upon Tyne, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.384807.
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Ferdous, Saba. "Structural and conformational analysis of B-cell epitopes : component to guide peptide vaccine design." Thesis, University College London (University of London), 2018. http://discovery.ucl.ac.uk/10042240/.
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Peptide vaccines have many potential advantages including low cost, lack of need for cold-chain storage and safety. However, it is well known that approximately 90% of B-cell Epitopes (BCEs) are discontinuous in nature making it difficult to mimic them for creating vaccines. To perform a detailed structural analysis of these epitopes, they needs to be mapped onto antigen structures that are complexed with antibody. In order to obtain a clean dataset of antibody-antigen complex crystal structures, a pipeline was designed to process automatically and clean the antibody related structures from the PDB. To store this processed antibody structural data, a database (AbDb) was built and made available online. The degree of discontinuity in B-cell epitopes and their conformational nature was studied by mapping epitopes in the antibody-antigen dataset. The discontinuity of B-cell epitopes was analysed by defining extended ‘regions’ (R, consisting of at least 3 antibody-contacting residues each separated by ≤ 3 residues) and small fragments (F, antibody-contacting residues that do not satisfy the requirements for a region). Secondly, an algorithm was developed to classify region shape as linear, curved or folded. Molecular dynamics simulations were carried out on isolated epitope regions (wild type and mutant peptides). The mutant peptides have been designed by mutating non-contacting and hydrophobic residues in epitopes. Two types of mutations (hy- drophobic to alanine and hydrophobic to glutamine) have been studied using molec- ular dynamics simulations. Furthermore, the effect of end-capping on wild type and mutant epitope regions has been studied. Simulation studies were carried out on 5 linear and 5 folded shape regions. Out of these, 2 epitopes (one linear and one folded), along with their mutants and derivatives, were tested experimentally for conformational stability by CD spectroscopy and NMR. The binding of isolated epitopes with antibody was also validated by ELISA and SPR.
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McGowan, Jean Louise. "Pseudomonas aeruginosa exotoxin A : immunochemical analysis of the proposed elongation factor 2 binding site and ADP-ribosyltransferase cross-reactive epitope /." The Ohio State University, 1991. http://rave.ohiolink.edu/etdc/view?acc_num=osu1487688973682985.
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Mathews, James H. "Analysis of T-helper cell epitopes on the structural glycoproteins of Venezuelan equine encephalitis virus." Thesis, University of Surrey, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.334738.
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Fallahi, Firouzeh. "Characterization of epitopes on the rabies virus glycoprotein by selection and analysis of escape mutants." Thesis, University of Ottawa (Canada), 2005. http://hdl.handle.net/10393/26898.
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Rabies virus, a negative sense single-stranded RNA virus, is the type member of the genus Lyssavirus of the Rhabdoviridae . The glycoprotein (G), which projects from the surface of the lyssavirus particle, is the only protein known to be capable of eliciting the production of neutralizing antibodies and knowledge of the antigenic nature of this protein is therefore important. Five different antigenic sites have been mapped on the G protein. In this study, the isolation of mutants resisting antibody neutralization (escape mutants) was attempted by a selection strategy employing three distinct strains of rabies: Evelyn Rokitnicki Abelseth (ERA), Big Brown Bat (BBB), and Silver Haired Bat (SHB). No escape mutants were generated from BBB and SHB but a total of seven independent ERA mutants were recovered using monoclonal antibodies (Mabs) directed against antigenic sites I and IIIa of the glycoprotein. (Abstract shortened by UMI.)
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Nason, Emma L. "Structural analysis of BTV VP7 epitopes with regard to the location of putative cell binding sites." Thesis, University of Oxford, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.390528.
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Leitch, Robert A. "Analysis of immunodominant epitopes of Neisseria gonorrhoeae lipopolysaccharide using a competitive inhibition enzyme linked immunosorbent assay." Thesis, University of Ottawa (Canada), 1985. http://hdl.handle.net/10393/4930.
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Kuoppala, Oskar. "Cavitation analysis on test rig. : An experimental and CFD study executed in collaboration with Epiroc AB." Thesis, Umeå universitet, Institutionen för tillämpad fysik och elektronik, 2021. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-188337.
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This master thesis project was done in collaboration with Epiroc Group Ab. Epiroc supplies high-quality drills of various types that can be used both above and below ground. A major problem in their percussive rock drills is that that cavitation is formed. Cavitation is a phenomenon that occurs when a fluid is subject to a sudden pressure drop. This pressure drop causes the liquid to vaporize and create gas bubbles. These gas bubbles will cause erosion to the walls when imploded. These cavitation damages lead to drills breaking and parts having to be replaced preserved. An experimental rig was used to create cavitation. From the experimental rig, it was possible to measure the hydraulic transients that are created when the valve was closed. In this study, we examined whether one can visually see these damages occurring inside the pipe on valve parts that are subjected to these cavitation damages. CFD simulations were used to re-create the closing of the valve in the experimental rig. By exporting pressure data from the experiments one could compare the numerical result to the experimental data. It was also investigated if it is possible to see some connection between the gas formation and the damages seen visually from the experimental part. For the simulation the realizable k − ε methods were implemented with enhanced wall treatment. The mixture model was used since we have a multi-phase flow. Some visual damages were recognized during the experiments. However, no distinguished pattern or specific areas was established. From the simulations, it could be determined that they generated gas when the valve was closed. However, the pressure transients could not be replicated in the numerical result.
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Nastke, Maria-Dorothea. "T-cell epitopes from viral and tumor associated antigens induction and analysis of antigen-specific T cells /." [S.l. : s.n.], 2005. http://www.bsz-bw.de/cgi-bin/xvms.cgi?SWB12168223.
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Mallon, Dermot Henry. "Structural and electrostatic analysis of HLA B cell epitopes : inferences on immunogenicity and prediction of humoral alloresponses." Thesis, University of Cambridge, 2018. https://www.repository.cam.ac.uk/handle/1810/271772.
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Despite the use of potent immunosuppressive agents, injury secondary to the alloimmune response principally directed against mismatched HLA antigens, remains a significant cause of renal transplant dysfunction and loss. One of the principal ways to limit the detrimental effect of immune alloreactivity towards the graft is through minimisation of donor-recipient HLA mismatches, but this has not changed conceptually since the early period of clinical transplantation. The central hypothesis of the research described in this thesis is that assessment of histocompatibility based on evaluation of structural differences between donor and recipient HLA molecules has the potential to improve graft outcomes after kidney transplantation. Amino acid sequence analyses have been central to gaining an understanding of the relative immunogenicity of a mismatched donor HLA antigen in the context of a recipient's HLA repertoire. While these methods offer advantages over basic enumeration of HLA mismatches, regardless of how sophisticated they may be, they are unable to fully describe the intricate nature of the B cell epitope--BCR interaction that initiates a humoral response. This interaction requires, but is not limited to, structural and electrostatic potential complementarity. The aim of this thesis is to describe a fully structural description of HLA immunogenicity through analysis of the three-dimensional physicochemical environment at the surface of the HLA molecule. First, I describe the bioinformatic techniques that integrate HLA sequence and X-ray crystallographic data to produce accurate models of HLA molecular structure and, subsequently, calculate the electrostatic potential in the three-dimensional area surrounding the molecule. Through quantitative comparison of the three-dimensional electrostatic potential on the surface of each HLA molecule, Electrostatic Mismatch Score 3D (EMS-3D) is derived. EMS-3D represents the electrostatic disparity between a group of HLA molecules. Subsequently, I demonstrate, using examples of HLA mutagenesis studies described in the literature, that patterns of antigenicity are better explained through analysis of surface electrostatic potential than by analysis of sequence. Of greater clinical interest, the ability of EMS-3D to predict the development of \textit{de novo} alloantibody responses was assessed in a cohort of patients who received a standardised immunisation and had their antibody profile characterised using Luminex single-antigen beads. This analysis validated EMS-3D as a valuable predictor of the development and magnitude of an alloantibody response, and therefore as a potentially useful clinical tool in the allocation of donor organs. The clinical utility of EMS-3D was assessed in a national cohort of kidney transplants, which found EMS-3D to better predict transplant outcome than conventional, currently-employed, measures of histocompatibility. HLA polymorphisms are implicated in many disease processes, particularly in autoimmunity. In the last chapter, the methodology developed in this thesis is employed to analyse the association between particular HLA polymorphisms and Inflammatory Bowel Disease (IBD). Analyses of the electrostatic potential within the HLA peptide binding groove found that particular patterns of electrostatic potential within the HLA peptide binding groove were associated with disease phenotype. Future work will examine how the examination of multiple discrete regions, ostensibly epitopes, can further improve the prediction of the HLA alloimmune response.
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Michel, Yvonne [Verfasser], and Reinhard [Akademischer Betreuer] Bredehorst. "Structural and functional analyses of IgE epitopes and their biological relevance / Yvonne Michel. Betreuer: Reinhard Bredehorst." Hamburg : Staats- und Universitätsbibliothek Hamburg, 2012. http://d-nb.info/1024772802/34.
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Cossins, Judith Ann. "Prediction and analysis of murine K'krestricted cytotoxic cell epitopes in selected influenza A/PR/8/34 virus proteins." Thesis, University of Oxford, 1992. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.315710.
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Cherif, Alhaji. "Mathematical evolutionary epidemiology : limited epitopes, evolution of strain structures and age-specificity." Thesis, University of Oxford, 2015. http://ora.ox.ac.uk/objects/uuid:28dec0f4-e6da-466a-905c-d875f132415e.
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We investigate the biological constraints determined by the complex relationships between ecological and immunological processes of host-pathogen interactions, with emphasis on influenza viruses in human, which are responsible for a number of pandemics in the last 150 years. We begin by discussing prolegomenous reviews of historical perspectives on the use of theoretical modelling as a complementary tool in public health and epidemiology, current biological background motivating the objective of the thesis, and derivations of mathematical models of multi-locus-allele systems for infectious diseases with co-circulating serotypes. We provide detailed analysis of the multi-locus-allele model and its age-specific extension. In particular, we establish the necessary conditions for the local asymptotic stability of the steady states and the existence of oscillatory behaviours. For the age-structured model, results on the existence of a mild solution and stability conditions are presented. Numerical studies of various strain spaces show that the dynamic features are preserved. Specifically, we demonstrate that discrete antigenic forms of pathogens can exhibit three distinct dynamic features, where antigenic variants (i) fully self-organize and co-exist with no strain structure (NSS), (ii) sort themselves into discrete strain structure (DSS) with non-overlapping or minimally overlapping clusters under the principle of competitive exclusion, or (iii) exhibit cyclical strain structure (CSS) where dominant antigenic types are cyclically replaced with sharp epidemics dominated by (1) a single strain dominance with irregular emergence and re-emergence of certain pathogenic forms, (2) ordered alternating appearance of a single antigenic type in periodic or quasi-periodic form similar to periodic travelling waves, (3) erratic appearance and disappearance of synchrony between discrete antigenic types, and (4) phase-synchronization with uncorrelated amplitudes. These analyses allow us to gain insight into the age-specific immunological profile in order to untangle the effects of strain structures as captured by the clustering behaviours, and to provide public health implications. The age-structured model can be used to investigate the effect of age-specific targeting for public health purposes.
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Li, Anli. "The functional and structural analysis of integrin ߦ1 by mapping the epitopes of stimulatory mAbs of integrin ߦ1." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp05/mq23383.pdf.
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Dufour, Sophie. "Production of monospecific antibody to immunodominant epitopes of the caprine arthritis encephalitis virus transmembrane glycoprotein and analysis of their activity in vitro /." [S.l.] : [s.n.], 1999. http://www.stub.unibe.ch/html/haupt/datenbanken/diss/bestell.html.
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Popp, Jasmin [Verfasser], Harald [Akademischer Betreuer] Kolmar, and Thomas [Akademischer Betreuer] Holzhauser. "Analyses of IgE-epitope profiles from legume allergens as approach towards the development of novel diagnostic and therapeutic reagents in food allergy / Jasmin Popp ; Harald Kolmar, Thomas Holzhauser." Darmstadt : Universitäts- und Landesbibliothek Darmstadt, 2019. http://d-nb.info/1201820618/34.
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Eskilsson, Niklas, and David Magnuson. "Åtgärder för en effektivare intern materialförsörjning : Genomlysning av förbättringsområden för lager till slutmontering av gruvmaskiner med fördjupning inom frekvensläggning – en studie vid Epiroc Rock Drills AB." Thesis, Linköpings universitet, Logistik- och kvalitetsutveckling, 2019. http://urn.kb.se/resolve?urn=urn:nbn:se:liu:diva-157591.
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Epiroc har åtnjutit en längre period av kraftig tillväxt samtidigt som flertalet effektiviseringsprojekt har genomförts med syfte att öka produktionsvolymen för att möta marknadens efterfrågan. Detta genom att bland annat implementera en variant av Lean production – The Way We Produce. En av dessa förändringar är en takad flödesorienterad montering med just-in-time sekvenserad materialförsörjning. Detta har i sin tur ökat kraven på materialförrådet Logistikcenter (LC) där ledningen nu börjat undersöka möjliga effektiviseringsåtgärder. Därav är studiens syfte till att ta fram realiserbara förbättringsförslag för logistikverksamheten vid Logistikcenter tillhörande Epiroc Rock Drills AB i Örebro för att öka effektiviteten och leveranssäkerheten. Studien har genomförts i två faser; identifieringsfasen och fördjupningsfasen. Under identifieringsfasen genomfördes en kartläggning av nuläget i LC, där underlaget för kartläggningen baserar sig på intervjuer, observationer och analyser. Genom en rotorsaksanalys, med målet att identifiera källor till ineffektivitet, kunde nio förbättringsområden identifieras varav en av dessa vidare skulle utredas i fördjupningsfasen. Dessa utvärderades utifrån en effekt-insats matris för att välja det förbättringsområde med störst effektiviseringspotential i förhållande till den förväntande insatsen. Analysmodellen för effekt-insats matrisen var de åtta slöserierna i Lean (Petersson, et al., 2015), dess förväntade påverkan på effektiviteten samt den förväntade komplexiteten av en implementation. Resultatet från rotorsaksanalysen gav artikelklassificering som det primära förbättringsområdet där den undersökta åtgärden var en alternativ tillämpning av frekvensläggning för att minimera rörelsetiden mellan lagerplatser vid plock. Under fördjupningsfasen undersöktes den nuvarande artikelklassificeringen och frekvensläggningen genom syntes av en alternativ modell för klassificeringen av artiklar och lagerplatser utifrån en fördjupad litteraturstudie. För att undersöka om en alternativ klassificering kan öka effektiviteten utvecklades en utvärderingsmodell som modellerar rörelsetiden för historiskt data från plocklistor. Den användes för att testa vilken kombination av storlekar på artikelklasserna som gav den minsta möjliga totala rörelsetiden. En kombination av 60/30/10 % (A/B/C) av det ackumulerade antalet plock gav den lägsta totala rörelsetiden för en plockhistorik på 15 månader med en reduktion av rörelsetiden motsvarande 33 % (1760 h) jämfört mot nuläget. Vidare undersöktes en alternativ sortering av plocklistorna för automathissar respektive pallställage med utvärderingsmodellen som gav en reduktion på 4 % respektive 11 % mot nuläget. Slutligen undersöktes olika former på zonerna för klassificering av lagerplatser i pallställage W3, där utlämningsplatsen är placerad halvvägs in i ställaget. En tyngdpunkt placerat centralt mellan ingången och utlämningsplatsen gav det bästa simuleringsresultatet. Sammanfattningsvis fastställdes artikelklassificering som det förbättringsområde med störst realiserbar effektiviseringspotential med lägst komplexitet utifrån en rotsorsaksanalys. Epiroc rekommenderas att implementera klassificeringsmodellen med tre klasser av storleken 60/30/10 % (A/B/C) av den ackumulerade antalet plockrader, samt att implementera att den alternativa sorteringen av plockrader på plocklistor för pallställage.<br>Epiroc has had a long period of strong growth, where several efficiency projects have been implemented to increase production volume in order to meet market demand. A variant of Lean production has been implemented over several years – called The Way We Produce by Epiroc. A large part of that change has been the implementation of sequenced flow-oriented assembly with just-in-time sequenced material deliveries to the assembly floor. This, in turn, has increased the requirements for the warehouse Logistic Center (LC) and management has now begun to investigate ways to improve efficiency at LC. Hence, the aim of the study is to develop realistic improvement proposals for the logistics operations at Logistics Center of Epiroc Rock Drills AB at Örebro to increase efficiency and delivery reliability. The study has been conducted in two phases; the identification phase and the in-depth phase. During the identification phase, a mapping of the current situation in LC was carried out, where the basis for the survey is based on interviews, observations and analyzes. Through a root cause analysis, with the goal of finding sources of inefficiency, nine areas of improvement could be identified, one of which would be chosen for further investigation in the in-depth phase. The areas of improvement were evaluated with an effect-input matrix to choose the area of improvement that provides the greatest efficiency potential in relation to the expected effort. The analysis model for the effect-input matrix was the eight wastes of Lean based on Petersson et al. (2015), its expected impact on efficiency and the expected complexity of an implementation. The result of the root cause analysis gave article classification as the primary area of improvement, where the measure is an alternative slotting strategy to minimize the movement time between storage locations. During the in-depth phase, the current article classification and frequency setting were examined by synthesis of an alternative model for the classification of articles and storage locations based on an in-depth literature study. To investigate whether an alternative classification can increase efficiency, an evaluation model was developed that models the movement time from historical data from pick lists. It was used to test which combination of sizes for the article classes gave the smallest possible total movement time. A combination of 60/30/10% (A/B/C) gave the lowest total movement time for a picking history of 15 months with a reduction corresponding to 33% (1760 h) compared to the current situation. Furthermore, an alternative sorting of the pick lists for the vertical lift modules and pallet racking was examined with the evaluation model, which gave a reduction of 4% and 11% respectively. Finally, various forms for the zones were examined for the classification of storage locations in pallet rack W3, where the delivery site is located three-quarter way into the pallet rack. A center of gravity for the for the A-class placed between the entrance and the delivery point gave the best simulation result. In summary, the article classification improvement area was established with the greatest realizable efficiency potential with the least effort based on the root cause analysis. Epiroc is recommended to implement the classification model with three classes of size 60/30/10 % (A/B/C) of the accumulated number of picking rows, and to implement that alternative sorting of pick rows on picklists for pallet racking.
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Nastke, Maria-Dorothea [Verfasser], and Hans-Georg [Akademischer Betreuer] Rammensee. "T-cell epitopes from viral and tumor associated antigens : induction and analysis of antigen-specific T cells / Maria-Dorothea Nastke ; Betreuer: Hans-Georg Rammensee." Tübingen : Universitätsbibliothek Tübingen, 2005. http://d-nb.info/1162199148/34.
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Reynolds, Patrisha. "Temporal trends in grave marker attributes an analysis of headstones in Florida." Honors in the Major Thesis, University of Central Florida, 2012. http://digital.library.ucf.edu/cdm/ref/collection/ETH/id/607.
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Grave markers reflect a wealth of information and collectively epitomize society's historic, social, and economic patterns over time. Despite an abundance of cemetery research in other parts of the country, little research has been undertaken to evaluate grave marker attributes in Florida. The purpose of this research was to determine how grave marker attributes have changed over time in north-central, central, and southeast Florida. Data were collected from ten cemeteries in five counties in Florida, representing the grave markers of over 1,100 individuals. Data collection involved visiting each cemetery, photographing markers, and cataloging grave marker attributes. Attributes analyzed included marker type, marker material, epitaphs, iconographic images, memorial photographs, footstones, and kerbs. A number of important trends were noted. Marker material exhibited the clearest example of a temporal trend, shifting over time from 73% marble to 73% granite. Marker type varied greatly from upright and flat ground markers to a variety of customized markers and vaults. Cultural differences were also noted with in-ground vaults dominating traditionally black cemeteries. There were clear differences in marker style between affluent and less affluent cemeteries, with numerous hand-cast cement markers observed in less prosperous areas. Furthermore, beginning in the early 1980's there is an increase in customized laser engraved markers. Overall, Florida's cemeteries offer a rich history of the state's mortuary practices and further research should be conducted to preserve this history.<br>B.S.<br>Bachelors<br>Sciences<br>Anthropology
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Berry, Michael. "Cloning and characterization of the human coronavirus NL63 nucleocapsid protein." Thesis, University of the Western Cape, 2011. http://etd.uwc.ac.za/index.php?module=etd&action=viewtitle&id=gen8Srv25Nme4_2481_1365583304.
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<p>The human coronavirus NL63 was discovered in 2004 by a team of researchers in Amsterdam. Since its discovery it has been shown to have worldwide spread and affects mainly children, aged 0-5 years old, the immunocompromised and the elderly. Infection with HCoV-NL63 commonly results in mild upper respiratory tract infections and presents as the common cold, with symptoms including fever, cough, sore throat and rhinorrhoea. Lower respiratory tract findings are less common but may develop into more serious complications including bronchiolitis, pneumonia and croup. The primary function of the HCoV-NL63 nucleocapsid (N) protein is the formation of theprotective ribonucleocapsid core. For this particle to assemble, the N-protein undergoes N-N dimerization and then interacts with viral RNA. Besides the primary structural role of the Nprotein, it is also understood to be involved in viral RNA transcription, translation and replication, including several other physiological functions. The N-protein is also highly antigenic and elicits a strong immune response in infected patients. For this reason the N-protein may serve as a target for the development of diagnostic assays. We have used bioinformatic analysis to analyze the HCoV-NL63 N-protein and compared it to coronavirus N-homologues. This bioinformatic analysis provided the data to generate recombinant clones for expression in a bacterial system. We constructed recombinant clones of the N-protein of SARS-CoV and HCoV-NL63 and synthesized truncated clones corresponding to the N- and C-terminal of the HCoV-NL63 N-protein. These heterologously expressed proteins will serve the basis for several post-expression studies including characterizing the immunogenic epitope of the N-protein as well identifying any antibody crossreactivity between coronavirus species.</p>
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Lima, Raquel Vaccari de. "O gênero de discurso epitáfio e a imagem do outro na memória social." Pontifícia Universidade Católica de São Paulo, 2016. https://tede2.pucsp.br/handle/handle/18909.
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Previous issue date: 2016-03-21<br>Coordenação de Aperfeiçoamento de Pessoal de Nível Superior<br>The present thesis discusses the form in which the discourses of epitaphs, in a socio-historically constructed Christian context, deny the finitude of spiritual life and thus promote an idealized image of the deceased in order to eternalize him in the social memory and to extol, as a practice of a religious habitus, a posthumous life in “Eternity”. In the light of this, we have set as an overall goal to analyze the epitaph discourse genre and its socio-historical conditions of production. We have started off with the hypothesis that the Christian man, aware of his death, comforts himself with the certainty of eternal life and yearns to preserve his memory in his social group. The belief in death as a passage to another life perpetuates in the discourse of epitaphs, whose scenography assists the human being to enter “Eternity” and to list the biographical virtues of the deceased, thus legitimizing the perpetuation of his memory. We have adopted the theoretical-methodological framework of Discourse Analysis, especially the categories: discourse genre, scenography, interdiscourse and discursive ethos, proposed by Dominique Maingueneau. To deal with the considerations about death and epitaphs, we have fallen back upon aspects of approaches from Anthropology, Philosophy, Theology and Psychoanalysis. The analyses undertaken in the present research have pointed to, in the workings of the discourses in epitaphs, the crossing of the discursive fields of death, religiosity, literature and history. We have also discovered that the scenographies that engender the discourses of epitaphs range from the promotion of a posthumous life to biography, in which the exemplary behavior of the deceased is extolled. Finally, the research has pointed out to the fact that the discursive ethos of the enunciator corresponds to the positive image of the good Christian<br>Esta tese trata da forma como os discursos dos epitáfios, num contexto cristão sócio-historicamente constituído, negam a finitude da vida espiritual e promovem uma imagem exemplar do morto a fim de eternizá-lo na memória social e exaltar, como prática de um habitus religioso, uma vida póstuma na “Eternidade”. O objetivo geral é analisar os gêneros de discurso epitáfio e suas condições sócio-históricas de produção. Partimos da hipótese segundo a qual o homem cristão, consciente de sua morte, consola-se com a certeza da vida eterna e anseia por manter viva sua lembrança em seu grupo social. A crença na morte como passagem para outra vida perpetua-se no discurso dos epitáfios, cuja cenografia auxilia o ser humano a ingressar na “Eternidade” e a biografar as virtudes do falecido, legitimando a perpetuação de sua memória. Adotamos o aparato teórico-metodológico da Análise do Discurso, principalmente as categorias: gênero de discurso, cenografia, interdiscurso e ethos discursivo, propostas por Dominique Maingueneau. Para as considerações acerca da morte e dos epitáfios, recorremos a aspectos de abordagens da Antropologia, da Filosofia, da Teologia e da Psicanálise. As análises empreendidas nesta pesquisa apontaram, no funcionamento dos discursos dos epitáfios, o cruzamento de campos discursivos da morte, da religiosidade, do literário e o da história. Constatamos, ainda, que as cenografias que engendram os discursos dos epitáfios variam de forma ora de promoção de uma vida póstuma, ora biográfica, exaltando as condutas exemplares do morto. E, por fim, a pesquisa constatou que o ethos discursivo do enunciador é correlato à imagem positiva do bom cristão
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Hoff, Merle. "Kombinatorische Analyse von Nanobody-markierten Epitopen zur Proteinbestimmung." Doctoral thesis, 2021. http://hdl.handle.net/21.11130/00-1735-0000-0005-1565-2.
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Lo, Ying-Tsang, and 羅英倉. "Antigen binding surface patch analysis and conformational epitope prediction." Thesis, 2017. http://ndltd.ncl.edu.tw/handle/58ejbc.
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博士<br>國立臺灣海洋大學<br>資訊工程學系<br>105<br>It is yet a challenging task to analyze and predict binding regions between proteins and specific molecular recognition elements like ligands or peptides based on three dimension structural information. Because of heavy computational cost, most biologists selected alternative ways to discover protein binding situations through sequence analysis or biological experiments. Owing to advances in computer technology, the hardware ability and storage capacity are greatly enhanced, and it can significantly improve the bottleneck of computing resources and facilitate to analyze the biological nature from structural and conformational characteristics. In this thesis, we proposed an approach for predicting epitope-paratope binding regions. These techniques are important to the preprocessing procedures of drug design, and the predicted binding regions can be evaluated in both vaccine development and monoclonal antibody (mAb)-resistant variant studies.
A conformational epitope (CE) in an antigenic protein is composed of specific amino acid residues. CEs bind their complementary paratopes in B-cell receptors and/or antibodies. An effective and efficient prediction tool for epitope analysis plays an important role for growth and development in immune-related applications, such as vaccine design and disease diagnosis. In this research topic, a new conformational epitope region search and prediction system was proposed. The proposed system is divided into two sequential modules - search and prediction modules. Search module includes two approaches. The first Complete Sequence Search (CSS) method applies BLAST tools to search all existing known antigen sequences. Those fragments with high epitope sequence identities can be detected and the predicted residues are annotated on the query structure. The second Spiral Vector Search (SVS) method adopts a new surface spiral feature vector for large-scale surface patch detection against a complete epitope database. A comprehensive testing dataset was generated for evaluation, and two searching methods can effectively identify all epitope regions. Regarding prediction modules, two systems were constructed. The first CEKEG system predicts conformational epitopes based on the knowledge-based energy and geometrical neighboring residue contents, and the second SFVP system adopts combinatorial features including amino acid contents and physicochemical characteristics to formulate corresponding geometric spiral vectors and compares with all spiral vectors from known conformational epitopes. The prediction results show the proposed methods perform better than all previously published systems in terms of sensitivity, specificity, positive predictive value and accuracy.
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Prokopová, Tereza. "In vitro test buněčné imunitní odpovědi pro diagnostiku Lymeské boreliózy." Master's thesis, 2017. http://www.nusl.cz/ntk/nusl-367822.
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Lyme borreliosis is a multisystemic disease affecting skin, joints, heart and central nervous system. The disease is caused by spirochetes of Borrelia burgdorferi sensu lato complex. These bacteria are spread by ticks of Ixodes genus. In 2016 there were almost 4,000 newly infected individuals reported in the Czech Republic. Contemporary serological diagnostics of Lyme borreliosis is not sensitive nor specific enough and does not even correlate with the pathology of the disease in the early or late phases. For the correct diagnosis of the disease it is necessary to detect the pathogen and its genotype. For this reason we had aimed at two goals. Through the digital droplet PCR (ddPCR) method we detected Borrelia-specific DNA and its genotype. The detection limit of borrelial DNA was set on gDNA samples isolated from the tick. Detection threshold for the initial amount of 1 ng of tick gDNA is at the range of 10-17 g of specific borrelial DNA. Borrelia spp. coinfection was detected in 5 out of 12 tested samples. The most frequent type was B. garinii which was detected in 5 samples. On the basis of published sequences for virulent factors we have designed specific primers in conserved regions of the genes flanking their variable segments to be PCR amplified. Gene variability will be monitored through...
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Šímová, Michaela. "Detekce povrchového fenotypu a chemosenzitivity buněk nádorů močového měchýře in vitro." Master's thesis, 2021. http://www.nusl.cz/ntk/nusl-438307.
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Tumor malignancies are the second leading cause of death worldwide. One of the reasons for the failure of oncological treatment are the uniformly set clinical guidelines, which neglect the effect of high intertumoral heterogeneity. The in vitro chemosensitivity and resistance (CSRA) assays allow for the stratification of patients prior to therapy. Therefore, the CSRA are a long-considered method for personalization of components of chemotherapy regime. Nevertheless, none of them is being routinely used in clinical practice. Certain chemotherapeutics used for their cytotoxic and cytostatic effect are also able to induce so-called immunogenic cell death (ICD) of tumor cells and activate an anti-tumor immune response. Monitoring of changes in the expression of molecules associated with the regulation of the innate immune system on the surface of dying tumor cells would enable to predict the patient's ability to respond to treatment involving modern immunotherapeutics. The feasibility of CSRA using flow cytometry and microscopy is critically evaluated in this thesis on a model of bladder cancer. Simultaneously, the correlation of the immunogenic phenotype of tumor cells and their sensitivity to selected chemotherapeutics is discussed.
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Husslik, Felix. "Analysis of the IgE epitope profile of the soybean allergen Gly m 4." Phd thesis, 2016. https://tuprints.ulb.tu-darmstadt.de/5736/1/Analysis%20of%20the%20IgE%20epitope%20profile%20of%20the%20soybean%20allergen%20Gly%20m%204_Dissertation_FelixHusslik_2016.pdf.
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Individuals with birch pollinosis may show allergic reactions after consumption of soybean-containing food. This is caused by cross-reaction of IgE directed against the major birch pollen allergen Bet v 1 with the structurally homologous allergen Gly m 4 from soybean. Hypersensitivity reactions in birch-soy allergy range from mild reactions to severe systemic reactions. Sera of birch pollen-allergic subjects may contain IgE to Gly m 4, even though no allergy to soy is present. Thus Gly m 4-specific IgE per se is not a suitable biomarker for birch-related soy allergy. To develop novel approaches for improved diagnosis and therapy of birch-soy allergy, knowledge on epitopes and IgE epitope profile of Gly m 4 for is needed. To date, data on epitopes of Bet v 1 and its homologous allergen Gly m 4 is very limited.
In this study, birch pollen-allergic patients with (27 subjects) and without (20 subjects) clinically confirmed allergy to soybean were included and analyzed regarding Gly m 4-specific serum IgE/IgG levels and epitope profiles of Gly m 4 for IgE. Specific IgE levels against Bet v 1, Gly m 4 and further soy allergens Gly m 5 and Gly m 6 were determined by ImmunoCAP™ and IgE binding to rGly m 4 and soy extract was tested in western blot. To analyze putative IgE epitopes of Gly m 4, non-allergenic Norcoclaurine synthase (NCS) from meadow rue was used as a model protein. NCS is structurally homologous to Gly m 4 but exhibits none to very little binding of Gly m 4-specific IgE antibodies enabling grafting of Gly m 4 epitopes onto the model protein. Potential candidate residues of Gly m 4 were selected by bioinformatic analysis of antibody-binding phage-displayed peptides and mapping of segments of Gly m 4 primary structure onto the molecular surface of the allergen. As a result a library of recombinant NCS variants with potential IgE binding was generated. In addition, five multiple substitutional variants of rGly m 4 with a total number of 18 amino acid substitutions crucial for IgE binding were generated and analyzed for antibody binding with patients’ sera. Furthermore a misfolded variant of each rBet v 1a and rGly m 4 was generated and defined molar ratios of folded/misfolded variants were compared in different immunological and physicochemical assays.
Gly m 4-specific median IgE and IgG levels of allergic (IgE: 9.3 kUA/L, IgG: 8.1 mgA/L) and non-allergic (IgE: 4.5 kUA/L, IgG: 8.3 mgA/L) subjects were comparable. The specific IgE levels did not correlate to the (severity of) clinical phenotypes. 51 candidate residues of Gly m 4 were selected for IgE epitope analysis and 46 potential functional IgE epitopes, single residues within a structural IgE epitope which dominate the energetics of allergen-IgE binding, were identified with IgE binding to ΔNCSN42/P49 variants. The putative functional IgE epitope pattern was individual for each patient and not distinguishable between allergic and non-allergic subjects. Using five rGly m 4 variants parts of the results of the NCS-based analyses could be confirmed with 18 potential functional IgE epitopes identified. 46 potential functional IgE epitopes clustered into six distinct putative IgE-binding areas on Gly m 4 and eleven NCS variants (ΔNCSN42/P49_1-11) presenting parts of these epitope areas were purified, showing a Gly m 4-type secondary structure according to CD measurements. Densitometric analysis for binding of IgE antibodies was performed via dot blot with sera of the study population but no characteristic IgE epitope pattern could be found. In contrast ΔNCSN42/P49_9 was identified as most suitable marker to distinguish soy allergic from tolerant patients in birch-related soybean allergy with a sensitivity and specificity of 68% and 93%, respectively. Using a pool of ΔNCSN42/P49 variants a depletion of Gly m 4-specific IgE binding to about 80% and 70% in pooled sera of patients with and without soybean allergy, respectively, was possible. Therefore identified putative functional IgE epitopes are specific for interaction with IgE antibodies in study population. With ΔNCSN42/P49_9 a differentiation between sensitization to Gly m 4 and clinically confirmed soybean allergy might be possible. The usage of a Gly m 4-specific epitope library might be a more promising tool for evaluating birch-related soybean allergy compared to ImmunoCAP™ and screening of substitutional Gly m 4 variants alone. However no correlations between patient’s IgE epitope profile and specific symptoms to soy or severity of allergic reactions could be found using NCS-based epitope library. Rather patient-specific epitope pattern might be relevant for birch-related soybean allergy. Therefore a thorough diagnosis by the combination of component-resolved diagnosis and profound epitope analysis might be mandatory in the future.
Using two misfolded variants of rBet v 1a and rGly m 4 the impact of unstructured allergens in rBet v 1a/rGly m 4 preparations was addressed with physico- and immunological assays. Both rBet v 1aS112P/R145P and rGly m 4S111P/L150P showed a highly disordered protein conformation and reduced IgE binding frequencies with analyzed patients’ sera. With CD spectroscopy, immunoblot, ELISA and RBL cell release assay defined combinations of native and unstructured allergen were assessed concerning secondary structure and IgE binding. Correlation of rBet v 1a content with secondary structure and IgE binding was suitable only at high rBet v 1aS112P/R145P levels in mixtures. CD spectroscopy and ELISA performed more precise compared to immunoblot and rat basophil cell mediator release assay where larger deviations between native and unstructured allergen were necessary. In addition, quantification of IgE-binding allergen was difficult for concentrations of rBet v 1a ≤10% in all assays. Overall, CD, ELISA and RBL cell release assay underestimated while immunoblot overestimated the actual level of rBet v 1a. Results of both misfolded variants rBet v 1aS112P/R145P and rGly m 4S111P/L150P might be used within screening of hypoallergenic molecules with potential use in treatment of allergies or in quality assessment of recombinant allergen preparations.
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Chiang, Chia-Hui, and 江家慧. "Analysis on the Epitaph of Ouyang Xiu." Thesis, 2015. http://ndltd.ncl.edu.tw/handle/75798541918244406846.
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Huang, Wei-fang, and 黃瑋芳. "Epitope Analysis and Immunological Studies of the ApfA Protein of Actinobacillus pleropnumoniae." Thesis, 2006. http://ndltd.ncl.edu.tw/handle/76013203874582698830.
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碩士<br>國立高雄大學<br>生物科技研究所<br>94<br>Actinobcacillus pleuropneumoniae, a Gram-negative bacterium, is the etiological agent of porcine pleuropneumonia and pulmonary haemorrhagic necrosis causing swine severe respiratory disease that leads to significant economic loss. The virulence factors of this pathogen have been identified including capsular polysaccharide, lipopolysaccharide, exotoxin, and outer membrane proteins. Generally speaking, the fimbriae are necessary for bacteria to colonize on the mucous membrane surface of the host. Therefore, the fimbriae of A. pleuropneumoniae should play an important role for infection. In this study, the structural protein of A. pleuropneumoniae fimbria, ApfA, was used to investigate the epitopes analysis and the potential for subunit vaccine development.
The apfA gene has been cloned into pQE30-UA vector, expressed in E. coli expression system and the expressed ApfA protein was purified. To analyze the epitopes of ApfA protein, we predicted the epitopes using an epitope analysis program. The results show that the most possible epitopes should locate at aa55-62, aa68-75, aa77-85, and aa131-138, respectively. The most hydrophilic region (aa 131- 138) of ApfA was synthesized an eight-mer peptide which conjugated with carrier protein multiple antigen peptides (MAPs) for further immunization analysis. Besides, we also created a serial of deleted apfA clones using ExoIII deletion method and expressed a serial of truncated proteins. The truncated ApfA proteins were detected with A. pleuropneumoniae serotype 1 polyclonal antiserum using Western blot analysis to predict the epitopes of ApfA. However, the data shown that there are cross-reactions between the antiserum and E. coli to interfere the epitopes analysis. These truncated proteins could be analyzed using monoclonal antibody in the future.
To understand the potential of ApfA as a subunit vaccine candidate, we used 50μg, 10μg recombinant ApfA protein, MAP-conjugated synthetic peptide as antigens to immunize mice. A commercial inactivated A. pleuropneumoniae vaccine and PBS also used as positive and negative controls. After second immunization, the titers of antisera increased significantly except PBS group. These mice were challenged with 5x108 colony-forming unit (cfu) of serotype 1 of A. pleuropneumoniae after 28 days of first immunization. The survival rate of 10μg ApfA and inactivated vaccine groups were 25% and other groups were 0% after 72 hours of virulence strain challenge. The growth inhibition test was performed using the antiserum of survival mice to incubate with A. pleuropneumoniae. The results shown that antiserum can inhibit the growth of this pathogen. In summary, the data reveal that ApfA can induce significant antibody titer which can inhibit the growth of A. pleuropneumoniae and the induced immune responses can provide protection for 10 ID50 of virulence strain challenge in mice. The recombinant ApfA protein should be used as a potential subunit vaccine candidate in the future.
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Lai, Jen-Feng, and 賴人鳳. "epitope analysis and immunogical studies of surface protein P42 of mycoplasma hyopneumoniae." Thesis, 2000. http://ndltd.ncl.edu.tw/handle/21871226778315360505.
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碩士<br>國立中山大學<br>生物科學系研究所<br>88<br>Mycoplasma hyopneumoniae causes swine enzootic pneumonia (SEP) and leads to economic loss worldwide. The mechanism of pathogenesis is still not clear. Since this pathogen remains extracellulary after infection, the surface proteins on M. hyopneumoniae should play very important roles in adhering and affecting tracheal mucosal cells. Therefore, the potential of using the surface proteins as the basic to develop molecular vaccine is currently being investigated.
The recombinant clone expressing the 42 kDa protein was isolated from the �� EMBL3 library and subjected to further studies. We cloned the antigen gene (P42) in an expression vector (pGEX4T-2) and expressed in E. coli (BL21). Then, the 42 kDa protein was purified to locate the epitopes using the following two methods: (1) Computer program (PC GENE) analysis: amino acids 321 to 327 of P42 were more hydrophilic than other regions. (2) Cleavage by chemicals: P42 was cleaved by hydroxylamine to three fragments, 3.8 kDa, 20.3 kDa and 16.4 kDa peptide (from N to C terminal). The epitope location was determined to be on the 16 kDa fragment using the rabbit anti-Mycoplasmsa hyopneumoniae antiserum.
Synthetic peptide (aa 321-327, conjugated with diphtheria toxoid) and the recombinant P42 protein (with or without adjuvants) were used as vaccine candidates to immunize animals. They all induce rather high immune response in mice. However, The synthetic peptide vaccine was observed to induce high titer of antibodies, but mainly on DT. The recombinant protein vaccine induced high level in humoral immune response, but low level in cellular immune response. Addition of adjuvant (TiterMaxâGold), enhances the production of both immune responses and produce higher IgG2a than IgG1. The additive of adjuvant (P42) also induce the experimental animals to express IL-2, IFN-g, and IL-4, however, mice immunized with recombinant P42 alone expresses only IL-4. The liquid culture growth inhibition tests revealed that the antiserum isolated from recombinant P42 immunized mice has greater inhibitory effect on the growth of mycoplasma than the antiserum isolated from mice immunized with the synthetic vaccine.
In conclusion, the P42 protein is part of P65, therefore is an incomplete HSP70. However, It is still a good immunogen. Therefore, it is proposed to include some immunostimulators (such as adjuvant, cytokine and T cell epitope) with the recombinant P42 in future vaccine development.
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Hsieh, Yao-ho, and 謝翱合. "B cell epitope mapping and analysis of HCMV pp65 antigen in SLE patients." Thesis, 2005. http://ndltd.ncl.edu.tw/handle/12043322921101846787.
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碩士<br>慈濟大學<br>分子生物及細胞生物研究所<br>93<br>Systemic lupus erythematosus (SLE) is a chronic inflammatory autoimmune disease. The exact cause of lupus remains a mystery, but it causes variable inflammatory destruction of skin、brain、heart、lungs、joints、blood-forming organs、kidneys and nervous system. The prevalence of SLE in Taiwan had been estimated as approximately fifty thousand (5/1000) predominantly female (male/female = 1/9) affected. In clinical diagnose, SLE patients after develop different kinds of autoantibodies, such as anti-double strand DNA (anti-dsDNA Ab)、anti-Smith antibody (anti-Sm Ab)、anti-small nuclear ribonucleoprotein antibody (anti-snRNP) and ect. Current investigations agreed that the mechanisms involved in promoting SLE including genetics (such as human leukocyte antigen (HLA) types)、complement deficiency、MHC、sex hormone、environmental factor (such as drug)、virus infection、ultraviolet light and stress.
Previous studies suggested that virus infection including Cytomegalovirus infection could increase the risk in the development of SLE. Cytomegalovirus infection is known to induce autoimmune diseases in mice. Our study focused on pp65 lower matrix phosphoprotein of HCMV (Human cytomegalovirus). The pp65 antigen has been reported to stimulate cell-mediated immunity by human and animal subjects following infection or immunization. However, the B cell epitope for pp65 remain elusive due to normal blood donors seldom develop antibody to it. We have revealed that adult SLE patients have elevated anti-pp65 activity. In order to verify the possible autoimmune associated B cell epitope, the pp65 antigen was divided into 501、510 and 672 nucleotide fragments, which expend the entire length of pp65 gene. Subsequently, these fragments were cloned into pGEX-KG and pET30 forming pp65-1、pp65-2、pp65-3 and pp65-F (full length) respectively. Expressed full-length pp65 and its fragments are examined against sera from SLE, none-SLE connective tissue disease patients (CTD), and normal control via western blotting. According to western blotting analysis the pp65-3 was recognized by SLE sera at a such higher frequency than pp65-1、2. Therefore, this study suggested that pp65-3 fragment contains B cell epitopes that could play a role in promoting SLE development.
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Liu, Han-lin, and 劉漢麟. "Epitope analysis and construction of a broad spectrum single chain Fv antibody against potyviruses." Thesis, 2009. http://ndltd.ncl.edu.tw/handle/01754119751161290946.
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碩士<br>國立臺灣大學<br>植物病理與微生物學研究所<br>97<br>Mixed infection of viral disease happens frequently in calla lily field. Detection methods simultaneously targeting multiple viruses should be developed in order to save the time and cost. We cloned and expressed the conserved region of the coat protein of calla lily-infecting potyviruses including Dasheen mosaic virus, Turnip mosaic virus, Zantedeschia mosaic virus and Zantedeschia mild mosaic virus as the antigen to produce the monoclonal antibody (mAb). A broad spectrum mAb (C4) against the potyviruses was screened. It could detect at least 10 potyviruses in addition to these four potyviruses. To clarify different binding ability of potyvirus to C4 mAb, phage display peptide library was used to determine the epitope reacting to C4 mAb. After three round of panning, 38 plaques were randomly selected for test and the supernatant of phage culture was analyzed by phage ELISA and competition ELISA. Twenty-three phage clones were sequenced and their sequences were aligned with those of five potyviruses. From the result of sequence alignment, there were two possible locations of the epitope (164 to 175 and 178 to 189). According to the Western analysis of virus-infected plant samples, the possibility of location 164 to 175 could be excluded. The result of phage Western blot indicated if the phage clone contained residue W, L, G, E/Q, V/I or Y/F, its affinity to C4 mAb increased. The amino acid sequence alignment of nine phage clones and potyviruses revealed that the epitope recognized by C4 mAb was WV(T)MMDGXXQV(I)EY(F). Furthermore, we constructed a broad spectrum C4 single chain Fv antibody fragment (C4 scFv) and used prokaryotic and eukaryotic expression systems to express the recombinant antibody. The variable regions of heavy chain and light chain which are 342 and 336 bp long were separately amplified by specific degenerate primers. The VH and VL fragments were assembled with linker DNA. The 30.26-kDa C4 scFv was expressed by pET29a(+) vector in E. coli BL21 (DE3) but it formed inclusion body. The refolded C4 scFv analyzed by ELISA and Western blot showed non-specific reactions to healthy plants. In order to avoid the problem of forming inclusion body, a Pichia expression system was used to express C4 scFv. The Pichia expression vector expressed a 29.73-kDa soluble scFv due to its secretion signal peptide. SDS-PAGE analysis showed the highest expression level of C4 scFv was at 96 hr after induced by 1% methanol. The specificity of the purified C4 scFv was analyzed by dot blot and Western blot. The result showed that C4 scFv could specifically bind to the epitope of potyvirus as C4 mAb. In the future, we will express C4 scFv in plants to evaluate its effect on potyvirus particle assembly.
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Chen, Yen-I. Grace 1977. "Proteomic analysis of the pre-mRNA splicing machinery utilizing chromosomal locus epitope tagging in metazoans." Thesis, 2007. http://hdl.handle.net/2152/3213.
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Epitope tagging in metazoans is an important tool for biochemical analyses and is generally accomplished by using trans-genes with in-frame epitope tags. However, protein levels from trans-genes are rarely representative of native levels. To overcome the shortcomings using trans-genes, epitope tags were introduced by homologous recombination technology, termed CLEP tagging (Chromosomal Locus EPitope tagging), immediately upstream of the stop codon of targeted genes in chicken B cell line DT40 and mouse embryonic stem (ES) cells. I first demonstrated the feasibility and promise of this technique in DT40 cells by purifying low abundance polypeptides and factors loosely associated with the SmD3 protein, a core protein participating in pre-mRNA splicing and mRNA turnover, with a TAP (tandem affinity purification) tag. Glycerol gradient separation was performed to further characterize the SmD3-associated protein complexes from the 200S fractions, corresponding to the supraspliceosomes. The purification included all five spliceosomal snRNAs. Most known snRNP-associated proteins, 5' end binding factors, 3' end processing factors, mRNA export factors, hnRNPs, and other RNA binding proteins were identified from the protein components. Intriguingly, the purified supraspliceosomes also contained a number of structural proteins, nucleoporins, chromatin remodeling factors, and several novel proteins that were absent from splicing complexes assembled in vitro. I showed that the in vivo analyses provide a more comprehensive list of polypeptides associated with pre-mRNA splicing apparatus as well as those that coupled transcription to the pre-mRNA processing steps. With similar techniques, the TAP tag was inserted into the chromosomal locus of a pre-mRNA splicing factor component, mSART-1 in live mice. Surprisingly, a profound autoimmune response was induced in homozygous-modified mice, due likely to an inappropriate stimulation of the immune system. I believe these mice will serve as a valuable tool for the studies of mammalian autoimmune diseases, especially those resulting from the generation of autoantibodies against RNP components.<br>text
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George, Jiya Marian. "Generation and analysis of mutated clonal scFv antibody fragments against R7V epitope of HIV-1." Diss., 2012. http://hdl.handle.net/2263/29320.
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Human immuno deficiency virus (HIV) incorporates host cellular protein, beta-2-microglobulin (β2m), into its surface envelope during budding. β2m is a cellular protein that belongs to the major histocompatibility complex (MHC) Class I molecules. Studies have shown anti- β2m monoclonal antibodies (mAbs) has the ability of to neutralize the virus. An epitope consisting of seven amino acids of the β2m protein designated as R7V produces antibodies that protect HIV infected people from progressing to AIDS. These protective antibodies, called anti-R7V antibodies, were able to neutralize different HIV isolates, despite their genomic variations, various cellular targets and geographic origin. Anti-R7V antibodies in the format of single chain variable fragments (scFvs) were produced in our laboratory using the M13 phage display technology. These scFv antibody fragments were used during in vitro studies for the detection and neutralization of the R7V antigen by enzyme linked immune sorbent assay (ELISA). The scFv fragments produced against the R7V epitope showed interaction, however the antibody-antigen affinity was too weak for the virus neutralization assay. Hence, this project focused on the affinity maturation of the anti-R7V scFv fragments through random mutagenesis using the error prone (EP) PCR method. The EP PCR method generated two mutated anti-R7V scFvs. The mutated clones were subcloned into the pAK400 expression vector. The computer-based models, created using the Swiss PDB Deep Viewer 4.02 software, were used to predict the antigen-binding site and affinity analysis of both parent and mutated scFv’s. Mutated clone 1 failed to bind to the R7V epitope whereas mutated clone 2 had similar binding pattern as the parent scFv. Mutated clone 2 was predicted to have a higher binding affinity compared to the parent scFv. The results obtained demonstrate the efficacy of EP PCR to generate high affinity antibodies. Future experiments using high affinity anti-R7V scFv’s may lead to its potential use in diagnostics, therapeutics or vaccine development. Copyright<br>Dissertation (MSc)--University of Pretoria, 2012.<br>Biochemistry<br>unrestricted
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Wang, Ping-Chieh, and 王炳傑. "Epitope Analysis and Immunological Studies of The Snake Venom of Cobra Through Phage Display Technology." Thesis, 2007. http://ndltd.ncl.edu.tw/handle/69557796783169840797.
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碩士<br>國立高雄大學<br>生物科技研究所<br>95<br>The way to deal with the snakebite patient is diagnosing rapidly who was bitten by what kind of poisonous snake and administrating the correct antivenin immediately. The preparation of antivenin was conducted by immunizing horses with detoxified snake venom, separating serum from collected horse blood, and purifying with ammonium sulfate. The titers of antisera during immunization period or final purified antivenins should be determined by enzyme linked immunosorbent assays (ELISAs) or venom neutralization tests. Both of these two methods need to use venom as antigen. However, poisonous snake is classified as preserve animal and it is difficult and dangerous to capture and breed snake and collect venom. Therefore, it is an important topic to develop a cheap, effective and easy-to-get venom substitute being used as antigen for relative assays. In this study, the phage display technology was used to search the crucial epitopes of snake venom and screen the epitope-carrying phages. It is expected that these phages can be used as venom substitute to resolve the snake venom hard-to-get problem and reduce the cost of assay.
Two phage displayed peptide libraries, ph.D.-7 and ph.D.c7c, were used to conduct the biopanning with antibodies of antivenin. Through the process of elution, amplification, and screening of three rounds biopanning, the number of high affinity phage population increases with the cycles of screening. The eluted phages number raised from 3.7x105 to 2.9x106 of the ph.D.-7 library, and 6x104 to 8x105 of the ph.D-c7c library, respectively. It indicates that the high affinity phage populations were selected through three times of biopanning.
Randomly select single phage from the third biopanning of ph.D.-7 and ph.D-c7c libraries, isolating phages DNA and sent to sequencing company for DNA sequence analysis. The results showed that the sequences of heptapeptides of SDPSSPS, MNFGMAG, TLGFIPW, MNFGFMA, and ALFGLPP were found in 5, 4, 3, 2, and 2 times of ph.D.-7 library. Alignment with the heptapeptide sequences of the selected phage clones, four consensus sequence -D P S S P S, M N F G-M A-, --G/L F I/V P W and A/M L FG/P L P- were found. The heptapeptide sequence KSSLLRN selected from ph.D.-c7c showed that conserve with the “KSSLL” sequence of Taiwan cobra (Naja Naja Atra) venom (Accession No.1CHV_S). Also, the heptapeptide sequence QADKHNK indicated that same “DKHN” sequence of Chinese cobra (Naja atra) cysteine-rich venom protein (Accession No. AY261468.1).
In the binding specificity test, the selected phages D23, D40, C2, C13, C14, C19, C29, C30, C31 and C33 can specifically bind to monovalent cobra antivenin IgG. The results show that these phages should carry the foreign amino acids corresponding to the epitopes of cobra snake venom. The results of dot blot indicated that these selected positive phages have the potential for serving as cobra venom substitute and developing diagnostic kit.
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LIN, YA-LUN, and 林亞倫. "Characterization of monoclonal antibodies against Japanese encephalitis virus E and NS1 protein and its epitope analysis." Thesis, 1991. http://ndltd.ncl.edu.tw/handle/69879136236137687113.
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Nacarino, Martínez Alejandra [Verfasser]. "Analysis of the influence of epitope flanking regions on MHC class I restricted antigen presentation / Alejandra Nacarino Martínez." 2007. http://d-nb.info/986990701/34.
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Lin, Shi-ting, and 林詩婷. "Epitope analysis on a differential protein P25 between Naja naja atra and Bungarus multicinctus venom using phage display technology." Thesis, 2009. http://ndltd.ncl.edu.tw/handle/4xc9gm.
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碩士<br>國立高雄大學<br>生物科技研究所<br>97<br>Poisonous snake bite is a global important issue. Several victims could not see or identify clearly what kind of the snake bit them. Therefore, it could reduce the misuse of antivenin and lower the occurrence of serum sickness through the rapid identification what kind of the snake bit them. This study analyzed epitopes on a differential protein that existed in different kinds of snake using phage display technology to facilitate the development of diagnostic reagent. Based on the analysis of protein electrophoresis, a differential 25-kDa protein, named P25, was found existing in Taiwan cobra Naja naja atra venom (NAV) but not in Bungarus multicinctus venom (BMV). Western blot analysis showed that P25 could be recognized by horse-derived anti-NAV antivenin. Therefore, P25 protein was used to study the differential epitopes between NAV and BMV. First of all, we recovered the P25 from protein electrophoresis gel and detoxified it with the treatment of 0.25% glutaraldehyde. Antiserum was prepared by rabbit immunized with 100 μg of detoxified P25 and the antiserum activity against P25 was verified by Western blot analysis. Anti-P25 IgG was purified through protein A, its activity verified again, and was used for biopanning with three different phage display peptide libraries. After three rounds of biopanning for each library, the eluted phage numbers (pfu/ml) increased from 2.29×107 to 4.48×108 in ph.D.-C7C library, 8.05×106 to 9.1×108 in ph.D.-7 library, and 7.75×107 to 3.69×109 in ph.D-12 library, respectively. It indicated that the high affinity phage clones were selected through three rounds of biopanning. Phage clones were randomly selected and sequenced, the deduced amino acid sequences were aligned and seven consensus sequences were found between them. Comparison with the sequences of known snake venoms in NCBI database, several sequences matched on the phospholipase A2 of Lapernis hardwickii (Accession No. AF144349.1) and cysteine-rich secretory protein (CRISP) of Naja naja atra (Accession No. Q7T1K6). Selected phage clones were further analyzed by dot blot assay using purified anti-P25 IgG. The results showed that the clones which has dot blot signal higher than positive control and no cross reaction with anti-BMV antiserum were phage C7-20 and D12-9. Competitive ELISA analysis revealed that the D12-9 clone could compete with P25 bound to anti-P25 IgG. The D12-9 sequence matched the amino acid residues Arg49 and Tyr52 on the active site of phospholipase A2 of Lapernis hardwickii. It indicated that the sequence of D12-9 has diagnostic potential in distinguishing the antivenin immunoreactivity
of NAV from BMV.