Higgledy-piggledy sets in projective spaces of small dimension

This work focuses on higgledy-piggledy sets of $k$-subspaces in $\text{PG}(N,q)$, i.e. sets of projective subspaces that are 'well-spread-out'. More precisely, the set of intersection points of these $k$-subspaces with any $(N-k)$-subspace $\kappa$ of $\text{PG}(N,q)$ spans $\kappa$ itself. We highlight three methods to construct small higgledy-piggledy sets of $k$-subspaces and discuss, for $k\in\{1,N-2\}$, 'optimal' sets that cover the smallest possible number of points. Furthermore, we investigate small non-trivial higgledy-piggledy sets in $\text{PG}(N,q)$, $N\leqslant5$. Our main result is the existence of six lines of $\text{PG}(4,q)$ in higgledy-piggledy arrangement, two of which intersect. Exploiting the construction methods mentioned above, we also show the existence of six planes of $\text{PG}(4,q)$ in higgledy-piggledy arrangement, two of which maximally intersect, as well as the existence of two higgledy-piggledy sets in $\text{PG}(5,q)$ consisting of eight planes and seven solids, respectively. Finally, we translate these geometrical results to a coding- and graph-theoretical context.Comment: [v1] 21 pages, 1 figure [v2] 21 pages, 1 figure: corrected minor details, updated bibliograph

The use of blocking sets in Galois geometries and in related research areas

Blocking sets play a central role in Galois geometries. Besides their intrinsic geometrical importance, the importance of blocking sets also arises from the use of blocking sets for the solution of many other geometrical problems, and problems in related research areas. This article focusses on these applications to motivate researchers to investigate blocking sets, and to motivate researchers to investigate the problems that can be solved by using blocking sets. By showing the many applications on blocking sets, we also wish to prove that researchers who improve results on blocking sets in fact open the door to improvements on the solution of many other problems

The role of LPCAT in regulating cellular inflammatory responses to toll-like receptor ligands

A novel lipid modifying enzyme, lysophosphatidylcholine acyltransferase (LPCAT), has been shown to regulate pro-inflammatory cytokine responses to lipopolysaccharide (LPS), a key molecule in initiating sepsis. Current research has shown that phospholipid metabolism may influence cell signalling, since that is dependent on the clustering of the receptor molecules into membrane microdomains (lipid rafts). LPCAT therefore, may be a target for novel anti-sepsis therapies. This study aimed to determine if LPCAT is essential in the regulation of pro-inflammatory cytokine responses to Gram-positive cell bacterial components, such as peptidoglycan (PG), lipoteichoic acid (LTA) and synthetic tripalmitoylated lipopeptide Pam3CSK4 in monocytes and lung epithelial cells. Inhibition of LPCAT activity consistently reduced TNF-α, IL-6 and IL-8 protein and mRNA levels in monocytes stimulated with all microbial ligands tested. Furthermore, cytokine production observed in monocytes primed with IFN-γ prior to ligand stimulation was twice that seen in unprimed cells, yet still inhibition of LPCAT significantly decreased the amplified inflammatory response. Lung epithelial cells, BEAS-2B, were more immunologically responsive to Pam3CSK4 where it consistently induced a high secretion of IL-8 and IL-6. However, whilst inhibition of LPCAT demonstrated a reduction in cytokine secretion, it was not a predominant as observed in monocytes indicating that LPCAT may have a lesser role in these cells.The secondary aim was to characterise the expression levels of 5 LPCAT enzymes in diverse cell types to ascertain if there is differential expression of the LPCATs, which may help to explain the altered effects that LPCAT inhibition produces on inflammatory cytokine production between cell types. Data in this report did not demonstrate one particular LPCAT iso-form to be highly expressed by monocytes, however literature has suggested that LPCAT2 might be the enzyme to modulate the inducible phospholipid remodeling pathways in innate immune cells. Further studies on inducible LPCAT expression in human cell lines is required to support these ideas.Previous studies have observed LPCAT to regulate translocation of TLR4 into membrane lipid raft domains and subsequent down-stream inflammatory responses. Although the translocation of TLR2 into membrane lipid rafts has been researched, its regulation by LPCAT has not, thus it was investigated in this study. Complications with experiments, however, did not allow this to be adequately assessed but, literature suggests this is the likely mechanism that LPCAT elicits its immuno-regulatory effect in response to microbial stimuli.The present study provided evidence that LPCAT influences the complex network of cell signalling involved in microbial responses, underlying it’s importance in inflammatory responses and potentially offering a target for novel anti-sepsis therapies

Linear codes meeting the Griesmer bound, minihypers and geometric applications

Coding theory and Galois geometries are two research areas which greatly influence each other. In this talk, we focus on the link between linear codes meeting the Griesmer bound and minihypers in finite projective spaces. Minihypers are particular (multiple) blocking sets. We present characterization results on minihypers, leading to equivalent characterization results on linear codes meeting the Griesmer bound. Next to being interesting from a coding-theoretical point of view, minihypers also are interesting for geometrical applications. We present results on maximal partial μ-spreads in PG(N, q), (μ + 1)|(N + 1), on minimal μ-covers in PG(N, q), (μ + 1)|(N + 1), on (N − 1)-covers of Q + (2N + 1, q), on partial ovoids and on partial spreads of finite classical polar spaces, and on partial ovoids of generalized hexagons, following from results on minihypers

Purification and Characterisation of Benzaldehyde Dehydrogenase I from Acinetobacter calcoaceticus and the TOL Plasmid Encoded Benzaldehyde Dehydrogenase and Benzyl Alcohol Dehydrogenase from Pseudomonas putida

1 Acinetobacter calcoaceticus can grow on either mandelate or benzyl alcohol as sole sources of carbon and energy. L(+)-Mandelate is metabolised to benzoate with phenylglyoxylate and benzaldehyde as intermediates, whereas benzyl alcohol is converted to benzoate with benzaldehyde as the only intermediate. Although the intermediates of the mandelate and benzyl alcohol pathways converge at the level of benzaldehyde, the enzymes of the pathways are quite separate because of the presence of two different benzaldehyde dehydrogenases. This thesis is concerned primarily with the purification and characterisation of benzaldehyde dehydrogenase I which is induced during growth on mandelate. This was intended to complete the characterisation of all of the enzymes of the mandelate and benzyl alcohol pathways in A. calcoaceticus. The project was subsequently enlarged to include the purification and characterisation of the benzaldehyde dehydrogenase and the benzyl alcohol dehydrogenase encoded by the TOL plasmid pWW53 in Pseudomonas putida MT53, because this would allow a more broadly-based comparison of five aldehyde and alcohol dehydrogenases, including a comparison of chromosomal and plasmid encoded enzymes

MicroCT analysis of connectivity in porous structures: optimising data acquisition and analytical methods in the context of tissue engineering

Micro-computed X-Ray Tomography (MicroCT) is one of the most powerful techniques available for the three dimensional characterisation of complex multi-phase or porous microarchitectures. The imaging and analysis of porous networks are of particular interest in tissue engineering due to the ability to predict various large-scale cellular phenomena through the micro-scale characterisation of the structure. However, optimising the parameters for MicroCT data capture and analyses requires a careful balance of feature resolution and computational constraints whilst ensuring that a structurally representative section is imaged and analysed. In this work, artificial datasets were used to evaluate the validity of current analytical methods by considering the effect of noise and pixel size arising from the data capture, and intrinsic structural anisotropy and heterogeneity. A novel ‘segmented percolation method’ was developed to exclude the effect of anomalous, non-representative features within the datasets, allowing for scale-invariant structural parameters to be obtained consistently and without manual intervention for the first time. Finally, an in-depth assessment of the imaging and analytical procedures are presented by considering percolation events such as micro-particle filtration and cell sieving within the context of tissue engineering. Along with the novel guidelines established for general pixel size selection for MicroCT, we also report our determination of 3 µm as the definitive pixel size for use in analysing connectivity for tissue engineering applications.MN was sponsored by Geitslich Pharma AG and the Gates Cambridge Trus

Simple polystyrene Microfluidic Device for Sensitive and Accurate SERS-Based Detection of Infection By Malaria Parasites

Early and accurate detection of infection by pathogenic microorganisms, such as Plasmodium, the causative agent of malaria, is critical for clinical diagnosis and ultimately determines the patient’s outcome. We have combined a polystyrene-based microfluidic device with an immunoassay which utilises Surface-Enhanced Raman Spectroscopy (SERS) to detect malaria. The method can be easily translated to a point-of-care testing format and shows excellent sensitivity and specificity, when compared to the gold standard for laboratorial detection of Plasmodium infections. The device can be fabricated in less than 30 min by direct patterning on shrinkable polystyrene sheets of adaptable three-dimensional microfluidic chips. To validate the microfluidic system, samples of P. falciparum-infected red blood cell cultures were used. The SERS-based immunoassay enabled the detection of 0.0012 ± 0.0001 % parasitaemia in a P. falciparum-infected red blood cell culture supernatant, an ~7-fold higher sensitivity than that attained by most rapid diagnostic tests. Our approach successfully overcomes the main challenges of the current Plasmodium detection methods, including increased reproducibility, sensitivity, and specificity. Furthermore, our system can be easily adapted for detection of other pathogens and has excellent properties for early diagnosis of infectious diseases, a decisive step towards lowering their high burden on healthcare systems worldwide

Manufacturing of human mesenchymal stem cells: the analytical challenges

It has been repeatedly proven that cell therapies can address many current unmet clinical treatment needs and also improve on current treatment options for various diseases, from neurological disorders to bone repair (Rosset et al. 2014; Corey et al. 2017). Though the potential of cell therapies has been demonstrated at a relatively small scale, the realisation of bringing cell based treatments to a larger market is hindered by the complexity of the product along with safety concerned and high production cost. Safety concerns can be informed with more in-depth analytical analysis of the product, however this in turn increase the costs involved in producing a cell therapy (Davie et al. 2012). Consequently the cost of analytical techniques also needs to be reduced, to address this need the area of microfluidic based bioanalytics holds much promise (Titmarsh et al. 2014). The culturing of human mesenchymal stem cells (hMSC) was used as a proof of concept model to demonstrate where improved bioanalytical and bioassay methods could be utilised in the production of cell therapies. Cells from four donors were cultured under three different oxygen environments and the conditioned medium assessed for pro-angiogenic capabilities using a tube formation bioassay and a proportion of the cytokine secretome profile measured using Luminex technology. Thorough secretome analysis it was shown that predicting cytokine levels based solely on the donor was not possible as the handling of the cells also had an influence on the secretome profile. The donor expression profiles did not behave in the same manner across all oxygen environments, for example in some donors IL-8 levels increased per cell at lower oxygen where as other donors showed a decrease per cell. While the tube formation assay showed some differences between donors in pro-angiogenic capabilities it also highlights the challenges with interpreting large data sets. The feasibility of using a microcapillary film (MCF) based enzyme-linked immunosorbent assay (ELISA) to detected two relevant cytokines, IL-8 and hepatocyte growth factor (HGF) was investigated. Following on from this work the development of a combined MCF ELISA assay with hMSC cell culture to produce a fully closed cell screening system was initiated. It was shown that it was feasible to measure IL-8 and HGF using the MCF ELISA platform but further work would need to be done to make the system more compatible with the manufacturing environment. In order to adapt the MCF to also be an hMSC culture platform the first challenge was to functionalise the Fluorinated Ethylene Propylene (FEP) surface of the MCF. It was concluded that a poly (vinyl- alcohol) (PVA) and gelatin mixture produced a homogenous coating to which a consistent level of hMSC would attach. This work was carried out on a flat surface; therefore steps were taken to adapt this knowledge into the MCF, while there was evidence of hMSCs present inside the MCF more work will need to be done to bring this concept to an established platform

The development of immunoassays for the detection of bovine brucellosis and aflatoxin B1

The research discussed in this thesis focuses on the development and characterisation of immunoassays for the detection of aflatoxin Bi (AFBi), a toxic fungal metabolite, and for the diagnosis of bovine brucellosis, a highly contagious disease of cattle caused by B ru c e lla abortus. An AFBi-specific single chain fragment variable (scFv) was isolated from a preimmunised phage display library and the gene encoding it sub-cloned into a range of different vectors for soluble expression o f monomeric, dimeric and bifunctional scFvs. The genetically-derived scFvs were then applied to the development of competitive ELIS As and BIAcore-based inhibition assays for the detection o f AFBi. A lateral flow immunoassay (LFIA) was developed for the detection of AFBi using an AFBi-specific monoclonal antibody. Each immunoassay format described was suitable for the detection of AFBi, with high levels of sensitivity, specificity and reproducibility achieved. Several immunoassay formats for the diagnosis of bovine brucellosis, in serum samples, were investigated. Four antigens were selected as diagnostic markers for brucellosis and included whole B. abortus cells, a crude cytoplasmic lysate, an 18kDa cytoplasmic protein ( p i8) and a 26kDa periplasmic protein (bp26). Recombinant forms of the p i 8 and bp26 proteins were cloned and expressed using a high-level expression vector in E.coli. Two polyclonal antibodies, directed against whole B.abortus cells and the crude cytoplasmic lysate, were developed and a naive phage display library was used to isolate scFvs directed against the recombinant bp26. Feasibility studies were carried out on the indirect ELISAs incorporating the four antigens and on the sandwich ELISAs with the polyclonal antibodies. The indirect and sandwich ELISAs, for the diagnosis of bovine brucellosis, were then validated using a panel of Brucella - positive and negative serum samples