Production and Evaluation of Antibodies and Phage Display-Derived Peptide Ligands for Immunomagnetic Separation of Mycobacterium bovis

This study describes the development and optimization of an immunomagnetic separation (IMS) method to isolate Mycobacterium bovis cells from lymph node tissues. Gamma-irradiated whole M. bovis AF2122/97 cells and ethanol-extracted surface antigens of such cells were used to produce M. bovis-speci?c polyclonal and monoclonal antibodies in rabbits and mice. They were also used to generate M. bovis-speci?c peptide ligands by phage display biopanning. The various antibodies and peptide ligands obtained were used to coat MyOne tosyl-activated Dynabeads (Life Technologies), singly or in combination, and evaluated for IMS. Initially, M. bovis capture from Middlebrook 7H9 broth suspensions (concentration range, 10 to 10⁵ CFU/ml) was evaluated by IMS combined with an M. bovis-speci?c touchdown PCR. IMS-PCR results and, subsequently, IMS-culture results indicated that the beads with greatest immunocapture capability for M. bovis in broth were those coated simultaneously with a monoclonal antibody and a biotinylated 12-mer peptide. These dually coated beads exhibited minimal capture (mean of 0.36% recovery) of 12 other Mycobacterium spp. occasionally encountered in veterinary tuberculosis (TB) diagnostic laboratories. When the optimized IMS method was applied to various M. bovis-spiked lymph node matrices, it demonstrated excellent detection sensitivities (50% limits of detection of 3.16 and 57.7 CFU/ml of lymph node tissue homogenate for IMS-PCR and IMS-culture, respectively). The optimized IMS method therefore has the potential to improve isolation of M. bovis from lymph nodes and hence the diagnosis of bovine tuberculosis.

Display; A Lie

Two original poems

Production and evaluation of the utility of novel phage display-derived peptide ligands to Salmonella spp. for magnetic separation

Aims: The objectives of this study were to produce Salmonella-specific peptide ligands by phage display biopanning and evaluate their use for magnetic separation (MS).
Methods and Results: Four phage display biopanning rounds were performed and the peptides expressed by the two most Salmonella-specific (on the basis of phage binding ELISA results) phage clones, MSal020401 and MSal020417, were chemically synthesized and coupled to MyOne™ tosylactivated Dynabeads®. Peptide capture capability for whole Salmonella cells from non-enriched broth cultures was quantified by MS + plate counts and MS + Greenlight™ detection, and compared to capture capability of anti-Salmonella (antibody-coated) Dynabeads®. MS + Greenlight™ gave a more comprehensive picture of capture capability than MS + plate counts and showed that Peptide MSal020417-coated beads exhibited at least similar, if not better, capture capability to anti-Salmonella Dynabeads® (mean capture values of 36.0 ± 18.2 % and 31.2 ± 20.1 %, respectively, over Salmonella spp. concentration range 3 x 101 - 3 x 106 cfu ml-1) with minimal cross-reactivity (= 1.9 %) to three other foodborne bacteria.
Conclusions: One of the phage display-derived peptide ligands was demonstrated by MS + Greenlight™ to be a viable antibody-alternative for MS of Salmonella spp.
Significance and Impact of Study: This study demonstrates an antibody-free approach to Salmonella detection and opens substantial possibilities for more rapid tests for this bacterium.

Phage Display-Derived Binders Able to Distinguish Listeria monocytogenes from Other Listeria Species

The objective of this study was to produce phage display-derived binders with the ability to distinguish Listeria monocytogenes from other Listeria spp., which may have potential utility to enhance detection of Listeria monocytogenes. To obtain binders with the desired binding specificity a series of surface and solution phage-display biopannings were performed. Initially, three rounds of surface biopanning against gamma-irradiated L. monocytogenes serovar 4b cells were performed followed by an additional surface biopanning round against L. monocytogenes 4b which included prior subtraction biopanning against gamma-irradiated L. innocua cells. In an attempt to further enhance binder specificity for L. monocytogenes 4b two rounds of solution biopanning were performed, both rounds included initial subtraction solution biopanning against L. innocua. Subsequent evaluations were performed on the phage clones by phage binding ELISA. All phage clones tested from the second round of solution biopanning had higher specificity for L. monocytogenes 4b than for L. innocua and three other foodborne pathogens (Salmonella spp., Escherichia coli and Campylobacter jejuni). Further evaluation with five other Listeria spp. revealed that one phage clone in particular, expressing peptide GRIADLPPLKPN, was highly specific for L. monocytogenes with at least 43-fold more binding capability to L. monocytogenes 4b than to any other Listeria sp. This proof-of-principle study demonstrates how a combination of surface, solution and subtractive biopanning was used to maximise binder specificity. L. monocytogenes-specific binders were obtained which could have potential application in novel detection tests for L. monocytogenes, benefiting both the food and medical industries. © 2013 Morton et al.

Content of Museum display and objects, Gozo

2007- present- National Museum of Archaeology, Gozo- text, display, academic input

Il Neolitico di Umbria - gallery, San Marco Gubbio.:Neolithic text and selection of material for display and interpretation

2009-10 Perugia – provision of material and data for gallery and information panels in Perugia Museo dei Provincia

Adventitial stem cells in vein grafts display multilineage potential that contributes to neointimal formation

This study was designed to carry out the characterization of stem cells within the adventitia and to elucidate their functional role in the pathogenesis of vein graft atherosclerosis.

How Does Justice Sound? Enunciative Modalities, Law’s Logos and Woman’s Voice

In the throes of her mimetic exposure of the lie of phallocratic discursive unity in 'Speculum of the Other Woman', Irigaray paused on the impossibility of woman’s voice and remarked that ‘it [was] still better to speak only in riddles, allusions, hints, parables.’ Even if asked to clarify a few points. Even if people plead that they just don’t understand. After all, she said, ‘they never have understood.’ (Irigaray 1985, 143).

That the law has never understood a uniquely feminine narrative is hardly controversial, but that this erasure continues to have real and substantive consequences for justice is a reality that feminists have been compelled to remain vigilant in exposing. How does the authority of the word compound law’s exclusionary matrix? How does law remain impervious to woman’s voice and how might it hear woman’s voice? Is there capacity for a dialogic engagement between woman, parler femme, and law?

This paper will explore these questions with particular reference to the experience of women testifying to trauma during the rape trial. It will argue that a logically linked historical genealogy can be traced through which law has come to posit itself as an originary discourse by which thinking is very much conflated with being, or in other terms, law is conflated with justice. This has consequences both for women’s capacity to speak or represent the harm of rape to law, but also for law’s ability to ‘hear’ woman’s voice and objectively adjudicate in cases of rape. It will suggest that justice requires law acknowledge the presence of two distinct and different subjects and that this must be done not only at the symbolic level but also at the level of the parole, syntax and discourse.

Decoding semantics across fMRI sessions with different stimulus modalities:a practical MVPA study

Both embodied and symbolic accounts of conceptual organization would predict partial sharing and partial differentiation between the neural activations seen for concepts activated via different stimulus modalities. But cross-participant and cross-session variability in BOLD activity patterns makes analyses of such patterns with MVPA methods challenging. Here, we examine the effect of cross-modal and individual variation on the machine learning analysis of fMRI data recorded during a word property generation task. We present the same set of living and non-living concepts (land-mammals, or work tools) to a cohort of Japanese participants in two sessions: the first using auditory presentation of spoken words; the second using visual presentation of words written in Japanese characters. Classification accuracies confirmed that these semantic categories could be detected in single trials, with within-session predictive accuracies of 80-90%. However cross-session prediction (learning from auditory-task data to classify data from the written-word-task, or vice versa) suffered from a performance penalty, achieving 65-75% (still individually significant at p « 0.05). We carried out several follow-on analyses to investigate the reason for this shortfall, concluding that distributional differences in neither time nor space alone could account for it. Rather, combined spatio-temporal patterns of activity need to be identified for successful cross-session learning, and this suggests that feature selection strategies could be modified to take advantage of this.

High-density lipoprotein subfractions display proatherogenic properties in overweight and obese children

BACKGROUND: In adults, obesity-driven inflammation can lead to increased cardiovascular disease (CVD). However, information regarding childhood obesity and its inflammatory sequelae is less well defined. Serum amyloid-A (SAA) is an inflammatory molecule that rapidly associates with high-density lipoproteins (HDLs) and renders them dysfunctional. Therefore, SAA may be a useful biomarker to identify increased CVD potential in overweight and obese children.

METHODS: Young Hearts 2000 is a cross-sectional cohort study in which 92 children who were obese were matched for age and sex with 92 overweight and 92 lean children. HDL2 and HDL3 (HDL2&3) were isolated from plasma by a three-step rapid-ultracentrifugation procedure. SAA was measured in serum and HDL2&3 by an enzyme-linked immunosorbent assay procedure, and the activities of cholesterol ester transfer protein (CETP) and lecithin cholesteryl acyltransferase (LCAT) were measured by fluorimetric assays.

RESULTS: Trends across the groups indicated that SAA increased in serum and HDL2&3 as BMI increased, as did HDL2-CETP and HDL2-LCAT activities.

CONCLUSION: These results have provided evidence that overweight and obese children are exposed to an inflammatory milieu that impacts the antiatherogenic properties of HDL and that could increase CVD risk. This supports the concept that it is important to target childhood obesity to help minimize future cardiovascular events.

Practice and Policy in the UK with Children and Young People who Display Harmful Sexual Behaviour: An analysis and critical review

This article considers the trajectory and effectiveness of policy, procedures and practice in the UK since the early 1990s in responding to young people who display problematic and harmful sexual behaviours. It draws on data from three publications in which research, policy and practice in the last 20 years have been reviewed. Key themes raised by Masson and Hackett are revisited including: denial and minimisation; terminology and categorisation; similarities with other young offenders; the child protection and youth justice systems; and assessment and interventions. The authors find that there is improvement in recognition of, and practice in response to, this group of young people, but good practice standards are inconsistently applied. With devolution of political powers, Scotland and Northern Ireland are now embarking on a more strategic response than England. The absence of a public debate and prioritising of primary prevention of child sexual abuse is noted.