N,N'-Carbonyldiimidazole-mediated functionalization of superparamagnetic nanoparticles as vaccine carrier

Particulates with specific sizes and characteristics can induce potent immune responses by promoting antigen uptake of appropriate immuno-stimulatory cell types. Magnetite (Fe3O4) nanoparticles have shown many potential bioapplications due to their biocompatibility and special characteristics. Here, superparamagnetic Fe3O4 nanoparticles (SPIONs) with high magnetization value (70emug-1) were stabilized with trisodium citrate and successfully conjugated with a model antigen (ovalbumin, OVA) via N,N'-carbonyldiimidazole (CDI) mediated reaction, to achieve a maximum conjugation capacity at approximately 13μgμm-2. It was shown that different mechanisms governed the interactions between the OVA molecules and magnetite nanoparticles at different pH conditions. We evaluated as-synthesized SPION against commercially available magnetite nanoparticles. The cytotoxicity of these nanoparticles was investigated using mammalian cells. The reported CDI-mediated reaction can be considered as a potential approach in conjugating biomolecules onto magnetite or other biodegradable nanoparticles for vaccine delivery.

Breaking the DNA-vaccine bottleneck

Monash University in Australia has developed a new approach towards DNA vaccine development that has the potential to cut the time it takes to produce a vaccine from up to nine months to four weeks or less. The university has designed and filed a patent on a commercially viable, single-stage technology for manufacturing DNA molecules. The technology was used to produce malaria and measles DNA vaccines, which were tested to be homogeneous supercoiled DNA, free from RNA and protein contaminations and meeting FDA regulatory standards for DNA vaccines. The technique is based on customized, smart, polymeric, monolithic adsorbents that can purify DNA very rapidly. The design criteria of solid-phase adsorbent include rapid adsorption and desorption kinetics, physical composition, and adequate selectivity , capacity and recovery. The new show technology significantly improved binding capacities, higher recovery, drastically reduced use of buffers and processing time, less clogging, and higher yields of DNA.

The suitability of DEAE-Cl active groups on customized poly(GMA-co-EDMA) continuous stationary phase for fast enzyme-free isolation of plasmid DNA

The creation of a commercially viable and a large-scale purification process for plasmid DNA (pDNA) production requires a whole-systems continuous or semi-continuous purification strategy employing optimised stationary adsorption phase(s) without the use of expensive and toxic chemicals, avian/bovine-derived enzymes and several built-in unit processes, thus affecting overall plasmid recovery, processing time and economics. Continuous stationary phases are known to offer fast separation due to their large pore diameter making large molecule pDNA easily accessible with limited mass transfer resistance even at high flow rates. A monolithic stationary sorbent was synthesised via free radical liquid porogenic polymerisation of ethylene glycol dimethacrylate (EDMA) and glycidyl methacrylate (GMA) with surface and pore characteristics tailored specifically for plasmid binding, retention and elution. The polymer was functionalised with an amine active group for anion-exchange purification of pDNA from cleared lysate obtained from E. coli DH5α-pUC19 pellets in RNase/protease-free process. Characterization of the resin showed a unique porous material with 70% of the pores sizes above 300 nm. The final product isolated from anion-exchange purification in only 5 min was pure and homogenous supercoiled pDNA with no gDNA, RNA and protein contamination as confirmed with DNA electrophoresis, restriction analysis and SDS page. The resin showed a maximum binding capacity of 15.2 mg/mL and this capacity persisted after several applications of the resin. This technique is cGMP compatible and commercially viable for rapid isolation of pDNA.

LacO-LacI interaction in affinity adsorption of plasmid DNA

Current approaches for purifying plasmids from bacterial production systems exploit the physiochemical properties of nucleic acids in non-specific capture systems. In this study, an affinity system for plasmid DNA (pDNA) purification has been developed utilizing the interaction between the lac operon (lacO) sequence contained in the pDNA and a 64mer synthetic peptide representing the DNA-binding domain of the lac repressor protein, LacI. Two plasmids were evaluated, the native pUC19 and pUC19 with dual lacO3/lacOs operators (pUC19lacO3/lacOs), where the lacOs operator is perfectly symmetrical. The DNA-protein affinity interaction was evaluated by surface plasmon resonance using a Biacore system. The affinity capture of DNA in a chromatography system was evaluated using LacI peptide that had been immobilized to Streamline™ adsorbent. The KD-values for double stranded DNA (dsDNA) fragments containing lacO1 and lacO3 and lacOs and lacO3 were 5.7 ± 0.3 × 10 -11 M and 4.1 ± 0.2 × 10-11 M respectively, which compare favorably with literature reports of 5 × 10-10 - 1 × 10-9 M for native laCO1 and 1-1.2 × 10-10 M for lacO1 in a saline buffer. Densitometric analysis of the gel bands from the affinity chromatography run clearly showed a significant preference for capture of the supercoiled fraction from the feed pDNA sample. The results indicate the feasibility of the affinity approach for pDNA capture and purification using native protein-DNA interaction.

Affinity adsorption of plasmid DNA

The development of a protein-mediated dual functional affinity adsorption of plasmid DNA is described in this work. The affinity ligand for the plasmid DNA comprises a fusion protein with glutathione-S-transferase (GST) as the fusion partner with a zinc finger protein. The protein ligand is first bound to the adsorbent by affinity interaction between the GST moeity and gluthathione that is covalently immobilized to the base matrix. The plasmid binding is then enabled via the zinc finger protein and a specific nucleotide sequence inserted into the DNA. At lower loadings, the binding of the DNA onto the Fractogel, Sepharose, and Streamline matrices was 0.0078 ± 0.0013, 0.0095 ± 0.0016, and 0.0080 ± 0.0006 mg, respectively, to 50 μL of adsorbent. At a higher DNA challenge, the corresponding amounts were 0.0179 ± 0.0043, 0.0219 ± 0.0035, and 0.0190 ± 0.0041 mg, respectively. The relatively constant amounts bound to the three adsorbents indicated that the large DNA molecule was unable to utilize the available zinc finger sites that were located in the internal pores and binding was largely a surface adsorption phenomenon. Utilization of the zinc finger binding sites was shown to be highest for the Fractogel adsorbent. The adsorbed material was eluted with reduced glutathione, and the eluted efficiency for the DNA was between 23% and 27%. The protein elution profile appeared to match the adsorption profiles with significantly higher recoveries of bound GST-zinc finger protein.

Reawakening reflective capacity in the psychotherapy of schizophrenia : a case study

Disturbed sense of self has long been identified as a common experience among people suffering with schizophrenia. More recently, metacognitive deficits have been found to be a stable and independent feature of schizophrenia that contributes to disturbed self-experience and impedes recovery. Individual psychotherapy designed to target poor metacognition has been shown to promote a more coherent sense of self and enhanced recovery in people with schizophrenia. We provide a report of a 2-year individual psychotherapy with a patient suffering with chronic schizophrenia. Progress was assessed over the course of treatment using the Metacognition Assessment Scale and the Brief Psychiatric Rating Scale. The patient experienced improved metacognitive capacity and reduced symptom severity over the course of therapy. Implications for clinical practice are discussed.

Multi-objective models and techniques for analysing the absolute capacity of railway networks

Railway capacity determination and expansion are very important topics. In prior research, the competition between different entities such as train services and train types, on different network corridors however have been ignored, poorly modelled, or else assumed to be static. In response, a comprehensive set of multi-objective models have been formulated in this article to perform a trade-off analysis. These models determine the total absolute capacity of railway networks as the most equitable solution according to a clearly defined set of competing objectives. The models also perform a sensitivity analysis of capacity with respect to those competing objectives. The models have been extensively tested on a case study and their significant worth is shown. The models were solved using a variety of techniques however an adaptive E constraint method was shown to be most superior. In order to identify only the best solution, a Simulated Annealing meta-heuristic was implemented and tested. However a linearization technique based upon separable programming was also developed and shown to be superior in terms of solution quality but far less in terms of computational time.

Capacity determination and expansion models for rail networks

This thesis focused upon the development of improved capacity analysis and capacity planning techniques for railways. A number of innovations were made and were tested on a case study of a real national railway. These techniques can reduce the time required to perform decision making activities that planners and managers need to perform. As all railways need to be expanded to meet increasing demands, the presumption that analytical capacity models can be used to identify how best to improve an existing network at least cost, was fully investigated. Track duplication was the mechanism used to expanding a network's capacity, and two variant capacity expansion models were formulated. Another outcome of this thesis is the development and validation of bi objective models for capacity analysis. These models regulate the competition for track access and perform a trade-off analysis. An opportunity to develop more general mulch-objective approaches was identified.

Modelling the BRT station capacity and queuing for all stopping busway operation

Stations on Bus Rapid Transit (BRT) lines ordinarily control line capacity because they act as bottlenecks. At stations with passing lanes, congestion may occur when buses maneuvering into and out of the platform stopping lane interfere with bus flow, or when a queue of buses forms upstream of the station blocking inflow. We contend that, as bus inflow to the station area approaches capacity, queuing will become excessive in a manner similar to operation of a minor movement on an unsignalized intersection. This analogy was used to treat BRT station operation and to analyze the relationship between station queuing and capacity. We conducted microscopic simulation to study and analyze operating characteristics of the station under near steady state conditions through output variables of capacity, degree of saturation and queuing. In the first of two stages, a mathematical model was developed for all stopping buses potential capacity with bus to bus interference and the model was validated. Secondly, a mathematical model was developed to estimate the relationship between average queue and degree of saturation and calibrated for a specified range of controlled scenarios of mean and coefficient of variation of dwell time.

Incorporating complex train paths to an analysis of absolute capacity

Railways are an important mode of transportation. They are however large and complex and their construction, management and operation is time consuming and costly. Evidently planning the current and future activities is vital. Part of that planning process is an analysis of capacity. To determine what volume of traffic can be achieved over time, a variety of railway capacity analysis techniques have been created. A generic analytical approach that incorporates more complex train paths however has yet to be provided. This article provides such an approach. This article extends a mathematical model for determining the theoretical capacity of a railway network. The main contribution of this paper is the modelling of more complex train paths whereby each section can be visited many times in the course of a train’s journey. Three variant models are formulated and then demonstrated in a case study. This article’s numerical investigations have successively shown the applicability of the proposed models and how they may be used to gain insights into system performance.

Watermarking capacity control for dynamic payload embedding

Despite significant improvements in capacity-distortion performance, a computationally efficient capacity control is still lacking in the recent watermarking schemes. In this paper, we propose an efficient capacity control framework to substantiate the notion of watermarking capacity control to be the process of maintaining “acceptable” distortion and running time, while attaining the required capacity. The necessary analysis and experimental results on the capacity control are reported to address practical aspects of the watermarking capacity problem, in dynamic (size) payload embedding.

The QUT Training Model for Peer Leader Capacity Building

QUT has enacted a university-wide Peer Program’s Strategy which aims to improve student success and graduate outcomes. A component of this strategy is a training model providing relevant, quality-assured and timely training for all students who take on leadership roles. The training model is designed to meet the needs of the growing scale and variety of peer programs, and to recognise the multiple roles and programs in which students may be involved during their peer leader journey. The model builds peer leader capacity by offering centralised, beginning and ongoing training modules, delivered by in-house providers, covering topics which prepare students to perform their role safely, inclusively, accountably and skilfully. The model also provides efficiencies by differentiating between ‘core competency' and ‘program-specific’ modules, thus avoiding training duplication across multiple programs, and enabling training to be individually and flexibly formatted to suit the specific and unique needs of each program.

The Yonder project: Stories of impact, change, and capacity building through engagement in arts education

This thesis examined the long-term impact of the community arts education project Yonder, a collaboration between Education Queensland and Queensland Performing Arts Centre. The findings from the data reveal that the project was still having impact twelve months after its completion and that in some instances the project served as a 'circuit-breaker', especially for special needs students and struggling students. The intervention of a rich arts project proved to be an opportunity for these students to learn in a different way and to perceive themselves in a new and reinvented light. This confidence was found to transfer into other aspects of their learning.

Medical education and law: Withholding/withdrawing treatment from adults without capacity

BACKGROUND Law is increasingly involved in clinical practice, particularly at the end of life, but undergraduate and postgraduate education in this area remains unsystematic. We hypothesised that attitudes to and knowledge of the law governing withholding/withdrawing treatment from adults without capacity (the WWLST law) would vary and demonstrate deficiencies among medical specialists. AIMS We investigated perspectives, knowledge and training of medical specialists in the three largest (populations and medical workforces) Australian states, concerning the WWLST law. METHODS Following expert legal review, specialist focus groups, pre-testing and piloting in each state, seven specialties involved with end-of-life care were surveyed, with a variety of statistical analyses applied to the responses. RESULTS Respondents supported the need to know and follow the law. There were mixed views about its helpfulness in medical decision-making. Over half the respondents conceded poor knowledge of the law; this was mirrored by critical gaps in knowledge that varied by specialty. There were relatively low but increasing rates of education from the undergraduate to continuing professional development (CPD) stages. Mean knowledge score did not vary significantly according to undergraduate or immediate postgraduate training, but CPD training, particularly if recent, resulted in greater knowledge. Case-based workshops were the preferred CPD instruction method. CONCLUSIONS Teaching of current and evolving law should be strengthened across all stages of medical education. This should improve understanding of the role of law, ameliorate ambivalence towards the law, and contribute to more informed deliberation about end-of-life issues with patients and families.

The peer leader capacity building model: A student learning journey

The benefits of peer leader experiences in building graduate skills and capabilities, is well documented and recognised in the higher education sector (Ender & Kay, 2001; Lindsey, Weiler, Zarich, Haddock, Krafchick, & Zimmerman, 2014; Shook & Keup, J., 2012). While benefits are acknowledged, responsibility for identifying, structuring and recording the learning experiences and learning outcomes is charged to the student. This poster describes a framework ‘The Peer Leader Capacity Building Model’ that purposefully structures the peer-leader’s learning journey providing: timely training, moments of critical reflection and goal setting. The model articulates the fundamental interplay of learning and peer leader service which forms the peer ‘learnership’. The journey begins with the ‘aspiration’ phase where students come to understand their leadership opportunities, progressing through ‘enabling’ and ‘mastering’ phases where students shape their learner-leader experience, and finally, to the ‘contributing graduate’ phase where students emerge as competent graduates able to confidently participate in their communities and workplaces. In shifting from a program centric approach that priorities the needs of the mentees, the Peer Leader Capacity Building Model focuses on the individual as a peer leader encouraging the student to shape their individual ‘learnscape’ through consciously navigating both their curricula and co-curricular learning experiences.