Dynamic, invivo, real-time detection of retinal oxidative status in a model of elevated intraocular pressure using a novel, reversibly responsive, profluorescent nitroxide probe

Changes to the redox status of biological systems have been implicated in the pathogenesis of a wide variety of disorders including cancer, Ischemia-reperfusion (I/R) injury and neurodegeneration. In times of metabolic stress e.g. ischaemia/reperfusion, reactive oxygen species (ROS) production overwhelms the intrinsic antioxidant capacity of the cell, damaging vital cellular components. The ability to quantify ROS changes in vivo, is therefore essential to understanding their biological role. Here we evaluate the suitability of a novel reversible profluorescent probe containing a redox-sensitive nitroxide moiety (methyl ester tetraethylrhodamine nitroxide, ME-TRN), as an in vivo, real-time reporter of retinal oxidative status. The reversible nature of the probe's response offers the unique advantage of being able to monitor redox changes in both oxidizing and reducing directions in real time. After intravitreal administration of the ME-TRN probe, we induced ROS production in rat retina using an established model of complete, acute retinal ischaemia followed by reperfusion. After restoration of blood flow, retinas were imaged using a Micron III rodent fundus fluorescence imaging system, to quantify the redox-response of the probe. Fluorescent intensity declined during the first 60 min of reperfusion. The ROS-induced change in probe fluorescence was ameliorated with the retinal antioxidant, lutein. Fluorescence intensity in non-Ischemia eyes did not change significantly. This new probe and imaging technology provide a reversible and real-time response to oxidative changes and may allow the in vivo testing of antioxidant therapies of potential benefit to a range of diseases linked to oxidative stress

Inertial sensor real-time feedback enhances the learning of cervical spine manipulation: A prospective study

Background Cervical Spinal Manipulation (CSM) is considered a high-level skill of the central nervous system because it requires bimanual coordinated rhythmical movements therefore necessitating training to achieve proficiency. The objective of the present study was to investigate the effect of real-time feedback on the performance of CSM. Methods Six postgraduate physiotherapy students attending a training workshop on Cervical Spine Manipulation Technique (CSMT) using inertial sensor derived real-time feedback participated in this study. The key variables were pre-manipulative position, angular displacement of the thrust and angular velocity of the thrust. Differences between variables before and after training were investigated using t-tests. Results There were no significant differences after training for the pre-manipulative position (rotation p = 0.549; side bending p = 0.312) or for thrust displacement (rotation p = 0.247; side bending p = 0.314). Thrust angular velocity demonstrated a significant difference following training for rotation (pre-training mean (sd) 48.9°/s (35.1); post-training mean (sd) 96.9°/s (53.9); p = 0.027) but not for side bending (p = 0.521). Conclusion Real-time feedback using an inertial sensor may be valuable in the development of specific manipulative skill. Future studies investigating manipulation could consider a randomized controlled trial using inertial sensor real time feedback compared to traditional training.

Real-time wind speed estimation and compensation for improved flight

This paper presents the development and experimental validation of a prototype system for online estimation and compensation of wind disturbances onboard small Rotorcraft unmanned aerial systems (RUAS). The proposed approach consists of integrating a small pitot-static system onboard the vehicle and using simple but effective algorithms for estimating the wind speed in real time. The baseline flight controller has been augmented with a feed-forward term to compensate for these wind disturbances, thereby improving the flight performance of small RUAS in windy conditions. The paper also investigates the use of online airspeed measurements in a closed-loop for controlling the RUAS forward motion without the aid of a global positioning system (GPS). The results of more than 80 flights with a RUAS confirm the validity of our approach.

Robust face clustering for real-world data

This thesis has investigated how to cluster a large number of faces within a multi-media corpus in the presence of large session variation. Quality metrics are used to select the best faces to represent a sequence of faces; and session variation modelling improves clustering performance in the presence of wide variations across videos. Findings from this thesis contribute to improving the performance of both face verification systems and the fully automated clustering of faces from a large video corpus.

Real-World Extensions To Scheduling Algorithms Based On Lagrangian Relaxation

Recently, efficient scheduling algorithms based on Lagrangian relaxation have been proposed for scheduling parallel machine systems and job shops. In this article, we develop real-world extensions to these scheduling methods. In the first part of the paper, we consider the problem of scheduling single operation jobs on parallel identical machines and extend the methodology to handle multiple classes of jobs, taking into account setup times and setup costs, The proposed methodology uses Lagrangian relaxation and simulated annealing in a hybrid framework, In the second part of the paper, we consider a Lagrangian relaxation based method for scheduling job shops and extend it to obtain a scheduling methodology for a real-world flexible manufacturing system with centralized material handling.

In situ reaction furnace for real-time XRD studies

The new furnace at the Materials Characterization by X-ray Diffraction beamline at Elettra has been designed for powder diffraction measurements at high temperature (up to 1373 K at the present state). Around the measurement region the geometry of the radiative heating element assures a negligible temperature gradient along the capillary and can accommodate either powder samples in capillary or small flat samples. A double capillary holder allows flow-through of gas in the inner sample capillary while the outer one serves as the reaction chamber. The furnace is coupled to a translating curved imaging-plate detector, allowing the collection of diffraction patterns up to 2[theta] [asymptotically equal to] 130°.

Real-time simulation of electrical machines on FPGA platform

This paper presents real-time simulation models of electrical machines on FPGA platform. Implementation of the real-time numerical integration methods with digital logic elements is discussed. Several numerical integrations are presented. A real-time simulation of DC machine is carried out on this FPGA platform and important transient results are presented. These results are compared to simulation results obtained through a commercial off-line simulation software.

Inhibition of 3-hydroxy-3-methylglutaryl coenzyme a reductase by cytosolic proteins - real or artifact

The inhibitory effect of FeSO4-dependent cytosolic protein on microsomal HMGCoA reductase is on the enzyme activity and not an artifact of loss of the product, mevalonate, through phosphorylation, unlike that of ATP.Mg effect.

Performance modelling of a fault-tolerant real-time multiprocessor using stochastic Petri nets

The fault-tolerant multiprocessor (ftmp) is a bus-based multiprocessor architecture with real-time and fault- tolerance features and is used in critical aerospace applications. A preliminary performance evaluation is of crucial importance in the design of such systems. In this paper, we review stochastic Petri nets (spn) and developspn-based performance models forftmp. These performance models enable efficient computation of important performance measures such as processing power, bus contention, bus utilization, and waiting times.

Towards real-time understanding of processes in pharmaceutical powder technology

There is a need for better understanding of the processes and new ideas to develop traditional pharmaceutical powder manufacturing procedures. Process analytical technology (PAT) has been developed to improve understanding of the processes and establish methods to monitor and control processes. The interest is in maintaining and even improving the whole manufacturing process and the final products at real-time. Process understanding can be a foundation for innovation and continuous improvement in pharmaceutical development and manufacturing. New methods are craved for to increase the quality and safety of the final products faster and more efficiently than ever before. The real-time process monitoring demands tools, which enable fast and noninvasive measurements with sufficient accuracy. Traditional quality control methods have been laborious and time consuming and they are performed off line i.e. the analysis has been removed from process area. Vibrational spectroscopic methods are responding this challenge and their utilisation have increased a lot during the past few years. In addition, other methods such as colour analysis can be utilised in noninvasive real-time process monitoring. In this study three pharmaceutical processes were investigated: drying, mixing and tabletting. In addition tablet properties were evaluated. Real-time monitoring was performed with NIR and Raman spectroscopies, colour analysis, particle size analysis and compression data during tabletting was evaluated using mathematical modelling. These methods were suitable for real-time monitoring of pharmaceutical unit operations and increase the knowledge of the critical parameters in the processes and the phenomena occurring during operations. They can improve our process understanding and therefore, finally, enhance the quality of final products.

Genetic variability of Tomato spotted wilt virus in Australia and validation of real time RT-PCR for its detection in single and bulked leaf samples

The potential for large-scale use of a sensitive real time reverse transcription polymerase chain reaction (RT-PCR) assay was evaluated for the detection of Tomato spotted wilt virus (TSWV) in single and bulked leaf samples by comparing its sensitivity with that of DAS-ELISA. Using total RNA extracted with RNeasy® or leaf soak methods, real time RT-PCR detected TSWV in all infected samples collected from 16 horticultural crop species (including flowers, herbs and vegetables), two arable crop species, and four weed species by both assays. In samples in which DAS-ELISA had previously detected TSWV, real time RT-PCR was effective at detecting it in leaf tissues of all 22 plant species tested at a wide range of concentrations. Bulk samples required more robust and extensive extraction methods with real time RT-PCR, but it generally detected one infected sample in 1000 uninfected ones. By contrast, ELISA was less sensitive when used to test bulked samples, once detecting up to 1 infected in 800 samples with pepper but never detecting more than 1 infected in 200 samples in tomato and lettuce. It was also less reliable than real time RT-PCR when used to test samples from parts of the leaf where the virus concentration was low. The genetic variability among Australian isolates of TSWV was small. Direct sequencing of a 587 bp region of the nucleoprotein gene (S RNA) of 29 isolates from diverse crops and geographical locations yielded a maximum of only 4.3% nucleotide sequence difference. Phylogenetic analysis revealed no obvious groupings of isolates according to geographic origin or host species. TSWV isolates, that break TSWV resistance genes in tomato or pepper did not differ significantly in the N gene region studied, indicating that a different region of the virus genome is responsible for this trait.

Detection of equine herpesvirus type 1 using a real-time polymerase chain reaction.

Equid herpesvirus 1 (EHV1) is a major disease of equids worldwide causing considerable losses to the horse industry. A variety of techniques, including PCR have been used to diagnose EHV1. Some of these PCRs were used in combination with other techniques such as restriction enzyme analysis (REA) or hybridisation, making them cumbersome for routine diagnostic testing and increasing the chances of cross-contamination. Furthermore, they involve the use of suspected carcinogens such as ethidium bromide and ultraviolet light. In this paper, we describe a real-time PCR, which uses minor groove-binding probe (MGB) technology for the diagnosis of EHV1. This technique does not require post-PCR manipulations thereby reducing the risk of cross-contamination. Most importantly, the technique is specific; it was able to differentiate EHV1 from the closely related member of the Alphaherpesvirinae, equid herpesvirus 4 (EHV4). It was not reactive with common opportunistic pathogens such as Escherichia coli, Klebsiella oxytoca, Pseudomonas aeruginosa and Enterobacter agglomerans often involved in abortion. Similarly, it did not react with equine pathogens such as Streptococcus equi, Streptococcus equisimilis, Streptococcus zooepidemicus, Taylorella equigenitalis and Rhodococcus equi, which also cause abortion. The results obtained with this technique agreed with results from published PCR methods. The assay was sensitive enough to detect EHV1 sequences in paraffin-embedded tissues and clinical samples. When compared to virus isolation, the test was more sensitive. This test will be useful for the routine diagnosis of EHV1 based on its specificity, sensitivity, ease of performance and rapidity.

Multiplex real-time PCR for the detection and differentiation of equid herpesvirus 1 (EHV-1) and equid herpesvirus 4 (EHV-4).

A multiplex real-time PCR was designed to detect and differentiate equid herpesvirus 1 (EHV-1) and equid herpesvirus 4 (EHV-4). The PCR targets the glycoprotein B gene of EHV-1 and EHV-4. Primers and probes were specific to each equine herpesvirus type and can be used in monoplex or multiplex PCRs, allowing the differentiation of these two closely related members of the Alphaherpesvirinae. The two probes were minor-groove binding probes (MGB?) labelled with 6-carboxy-fluorescein (FAM?) and VIC® for detection of EHV-1 and EHV-4, respectively. Ten EHV-1 isolates, six EHV-1 positive clinical samples, one EHV-1 reference strain (EHV-1.438/77), three EHV-4 positive clinical samples, two EHV-4 isolates and one EHV-4 reference strain (EHV-4 405/76) were included in this study. EHV-1 isolates, clinical samples and the reference strain reacted in the EHV-1 real-time PCR but not in the EHV-4 real-time PCR and similarly EHV-4 clinical samples, isolates and the reference strain were positive in the EHV-4 real-time PCR but not in the EHV-1 real-time PCR. Other herpesviruses, such as EHV-2, EHV-3 and EHV-5 were all negative when tested using the multiplex real-time PCR. When bacterial pathogens and opportunistic pathogens were tested in the multiplex real-time PCR they did not react with either system. The multiplex PCR was shown to be sensitive and specific and is a useful tool for detection and differentiation of EHV-1 and EHV-4 in a single reaction. A comprehensive equine herpesvirus disease investigation procedure used in our laboratory is also outlined. This procedure describes the combination of alphaherpesvirus multiplex real-time PCR along with existing gel-based PCRs described by other authors.

Absolute quantitation of Marek's disease virus and Herpesvirus of turkeys in chicken lymphocyte, feather tip and dust samples using real-time PCR

The further development of Taqman quantitative real-time PCR (qPCR) assays for the absolute quantitation of Marek's disease virus serotype 1 (MDV1) and Herpesvirus of turkeys (HVT) viruses is described and the sensitivity and reproducibility of each assay reported. Using plasmid DNA copies, the lower limit of detection was determined to be 5 copies for the MDV1 assay and 75 copies for the HVT assay. Both assays were found to be highly reproducible for Ct values and calculated copy numbers with mean intra- and inter-assay coefficients of variation being less than 5% for Ct and 20% for calculated copy number. The genome copy number of MDV1 and HVT viruses was quantified in PBL and feather tips from experimentally infected chickens, and field poultry dust samples. Parallelism was demonstrated between the plasmid-based standard curves, and standard curves derived from infected spleen material containing both viral and host DNA, allowing the latter to be used for absolute quantification. These methods should prove useful for the reliable differentiation and absolute quantitation of MDV1 and HVT viruses in a wide range of samples.

Embedding sustainability in a real-world context: Developing a world class early learning centre

This symposium describes what is possible when early childhood professionals work with designers to develop a vision for an exemplary early childhood centre with a focus on Education for Sustainability (EfS). The symposium provides insights into cross-disciplinary initiatives between QUT Early childhood and Design staff and students, who have worked together with the iconic Lone Pine Koala Sanctuary in Brisbane, to explore imperatives around EfS, including leadership and professionalism. This practical, real world project has seen all stakeholders engage in a focus on sustainability which has opened new ways of thinking about early childhood centre design. Cross-disciplinarity has created space to re-think the potential of the disciplines to interweave, and in so doing opened different ways for thinking about early childhood centres – their operation and their function. For the first time in Queensland, this project creates strategic alliances between EfS, childcare, business and sustainable design. EfS is essential for addressing local and global environmental issues and early childhood EfS research has been gaining international momentum, with governments nominating this area as having significant capacity to empower communities and promote change. While models for collaboration exist in the early childhood programs in Reggio Emilia, we offer sustainability as a unique and contemporary focus with immense potential to generate international and national interest. To date Early Childhood degree students enrolled in a leadership and management unit/subject have worked collaboratively with Design students to explore the sustainable design of the proposed Lone Pine early childhood centre. Providing students with a ‘real world’ project sees them re-positioned from ‘novice’ to ‘professional’, where their knowledge, expertise and perspectives are simultaneously validated and challenged. These learning experiences are enabling students to practice a new model of early childhood leadership, one that is vital for leading in an increasingly complex world. The symposium will be comprised of three discrete, though interconnected presentations, that work together to tell the story of this project. Three key facets of the project will be explored during the 90 minute session, as the perspectives of key stakeholders are shared. The first presentation (A/Prof Julie Davis, Dr Lyndal O’Gorman& Dr Megan Gibson) will outline the role of QUT School of Early Childhood staff and students, with attention to the ways in which the project was embedded in students’ work in the final year of their degree program of study. The second presentation (Ms Lindy Osborne) will provide insights into the Design students’ collaborative work in the project. Finally, the key role of the Lone Pine Koala Sanctuary and their commitment for EfS (Ms Peta Wilson & Dr Sue Elliott) will map out the philosophy that underpins the project. Together, the authors will conclude key project outcomes that have been achieved through this real-world, cross-disciplinary work.