Characterisation of a harvesting step for monoclonal antibodies from mammalian cell culture

The focus of this research was defined by a poorly characterised filtration train employed to clarify culture broth containing monoclonal antibodies secreted by GS-NSO cells: the filtration train blinded unpredictably and the ability of the positively charged filters to adsorb DNA from process material was unknown. To direct the development of an assay to quantify the ability of depth filters to adsorb DNA, the molecular weight of DNA from a large-scale, fed-batch, mammalian cell culture vessel was evaluated as process material passed through the initial stages of the purification scheme. High molecular weight DNA was substantially cleared from the broth after passage through a disc stack centrifuge and the remaining low molecular weight DNA was largely unaffected by passage through a series of depth filters and a sterilising grade membrane. Removal of high molecular weight DNA was shown to be coupled with clarification of the process stream. The DNA from cell culture supernatant showed a pattern of internucleosomal cleavage of chromatin when fractionated by electrophoresis but the presence of both necrotic and apoptotic cells throughout the fermentation meant that the origin of the fragmented DNA could not be unequivocally determined. An intercalating fluorochrome, PicoGreen, was elected for development of a suitable DNA assay because of its ability to respond to low molecular weight DNA. It was assessed for its ability to determine the concentration of DNA in clarified mammalian cell culture broths containing pertinent monoclonal antibodies. Fluorescent signal suppression was ameliorated by sample dilution or by performing the assay above the pI of secreted IgG. The source of fluorescence in clarified culture broth was validated by incubation with RNase A and DNase I. At least 89.0 % of fluorescence was attributable to nucleic acid and pre-digestion with RNase A was shown to be a requirement for successful quantification of DNA in such samples. Application of the fluorescence based assay resulted in characterisation of the physical parameters governing adsorption of DNA by various positively charged depth filters and membranes in test solutions and the DNA adsorption profile of the manufacturing scale filtration train. Buffers that reduced or neutralised the depth filter or membrane charge, and those that impeded hydrophobic interactions were shown to affect their operational capacity, demonstrating that DNA was adsorbed by a combination of electrostatic and hydrophobic interactions. Production-scale centrifugation of harvest broth containing therapeutic protein resulted in the reduction of total DNA in the process stream from 79.8 μg m1-1 to 9.3 μg m1-1 whereas the concentration of DNA in the supernatant of pre-and post-filtration samples had only marginally reduced DNA content: from 6.3 to 6.0 μg m1-1 respectively. Hence the filtration train was shown to ineffective in DNA removal. Historically, blinding of the depth filters had been unpredictable with data such as numbers of viable cells, non-viable cells, product titre, or process shape (batch, fed-batch, or draw and fill) failing to inform on the durability of depth filters in the harvest step. To investigate this, key fouling contaminants were identified by challenging depth filters with the same mass of one of the following: viable healthy cells, cells that had died by the process of apoptosis, and cells that had died through the process of necrosis. The pressure increase across a Cuno Zeta Plus 10SP depth filter was 2.8 and 16.5 times more sensitive to debris from apoptotic and necrotic cells respectively, when compared to viable cells. The condition of DNA released into the culture broth was assessed. Necrotic cells released predominantly high molecular weight DNA in contrast to apoptotic cells which released chiefly low molecular weight DNA. The blinding of the filters was found to be largely unaffected by variations in the particle size distribution of material in, and viscosity of, solutions with which they were challenged. The exceptional response of the depth filters to necrotic cells may suggest the cause of previously noted unpredictable filter blinding whereby a number of necrotic cells have a more significant impact on the life of a depth filter than a similar number of viable or apoptotic cells. In a final set of experiments the pressure drop caused by non-viable necrotic culture broths which had been treated with DNase I or benzonase was found to be smaller when compared to untreated broths: the abilities of the enzyme treated cultures to foul the depth filter were reduced by 70.4% and 75.4% respectively indicating the importance of DNA in the blinding of the depth filter studied.

Design of a stock control methodology for use within an oil blending company

The thesis deals with the background, development and description of a mathematical stock control methodology for use within an oil and chemical blending company, where demand and replenishment lead-times are generally non-stationary. The stock control model proper relies on, as input, adaptive forecasts of demand determined for an economical forecast/replenishment period precalculated on an individual stock-item basis. The control procedure is principally that of the continuous review, reorder level type, where the reorder level and reorder quantity 'float', that is, each changes in accordance with changes in demand. Two versions of the Methodology are presented; a cost minimisation version and a service level version. Realising the importance of demand forecasts, four recognised variations of the Trigg and Leach adaptive forecasting routine are examined. A fifth variation, developed, is proposed as part of the stock control methodology. The results of testing the cost minimisation version of the Methodology with historical data, by means of a computerised simulation, are presented together with a description of the simulation used. The performance of the Methodology is in addition compared favourably to a rule-of-thumb approach considered by the Company as an interim solution for reducing stack levels. The contribution of the work to the field of scientific stock control is felt to be significant for the following reasons:- (I) The Methodology is designed specifically for use with non-stationary demand and for this reason alone appears to be unique. (2) The Methodology is unique in its approach and the cost-minimisation version is shown to work successfully with the demand data presented. (3) The Methodology and the thesis as a whole fill an important gap between complex mathematical stock control theory and practical application. A brief description of a computerised order processing/stock monitoring system, designed and implemented as a pre-requisite for the Methodology's practical operation, is presented as an appendix.

High birefringence fibre interrogating interferometer for optical sensing applications

The use of high birefringence fiber interrogating interferometer for optical sensing applications was discussed. The method is of low cost and permits simple adjustment of the optical path difference and has much lower sensitivity to environmental perturbation. The polarization-maintaining (PM) fiber interferometer adopted a heterodyne approach using interferometric wavelength shift detection. The study showed that the inclusion of power amplifier driving a multi-element piezoelectric stack will enable the bandwidth to be pushed up into the kHz regime.

The factors affecting attitudes toward HSDPA service and intention to use:a cross-cultural comparison between Asia and Europe

HSDPA (High-Speed Downlink Packet Access) is a 3.5-generation asynchronous mobile communications service based on the third generation of W-CDMA. In Korea, it is mainly provided in through videophone service. Because of the diffusion of more powerful and diversified services, along with steep advances in mobile communications technology, consumers demand a wide range of choices. However, because of the variety of technologies, which tend to overflow the market regardless of consumer preferences, consumers feel increasingly confused. Therefore, we should not adopt strategies that focus only on developing new technology on the assumption that new technologies are next-generation projects. Instead, we should understand the process by which consumers accept new forms of technology and devise schemes to lower market entry barriers through strategies that enable developers to understand and provide what consumers really want.

Geospatial data quality indicators

Indicators which summarise the characteristics of spatiotemporal data coverages significantly simplify quality evaluation, decision making and justification processes by providing a number of quality cues that are easy to manage and avoiding information overflow. Criteria which are commonly prioritised in evaluating spatial data quality and assessing a dataset’s fitness for use include lineage, completeness, logical consistency, positional accuracy, temporal and attribute accuracy. However, user requirements may go far beyond these broadlyaccepted spatial quality metrics, to incorporate specific and complex factors which are less easily measured. This paper discusses the results of a study of high level user requirements in geospatial data selection and data quality evaluation. It reports on the geospatial data quality indicators which were identified as user priorities, and which can potentially be standardised to enable intercomparison of datasets against user requirements. We briefly describe the implications for tools and standards to support the communication and intercomparison of data quality, and the ways in which these can contribute to the generation of a GEO label.

UncertWeb processing service:making models easer to access on the web

Models are central tools for modern scientists and decision makers, and there are many existing frameworks to support their creation, execution and composition. Many frameworks are based on proprietary interfaces, and do not lend themselves to the integration of models from diverse disciplines. Web based systems, or systems based on web services, such as Taverna and Kepler, allow composition of models based on standard web service technologies. At the same time the Open Geospatial Consortium has been developing their own service stack, which includes the Web Processing Service, designed to facilitate the executing of geospatial processing - including complex environmental models. The current Open Geospatial Consortium service stack employs Extensible Markup Language as a default data exchange standard, and widely-used encodings such as JavaScript Object Notation can often only be used when incorporated with Extensible Markup Language. Similarly, no successful engagement of the Web Processing Service standard with the well-supported technologies of Simple Object Access Protocol and Web Services Description Language has been seen. In this paper we propose a pure Simple Object Access Protocol/Web Services Description Language processing service which addresses some of the issues with the Web Processing Service specication and brings us closer to achieving a degree of interoperability between geospatial models, and thus realising the vision of a useful 'model web'.

Discriminative training of the hidden vector state model for semantic parsing

In this paper, we discuss how discriminative training can be applied to the hidden vector state (HVS) model in different task domains. The HVS model is a discrete hidden Markov model (HMM) in which each HMM state represents the state of a push-down automaton with a finite stack size. In previous applications, maximum-likelihood estimation (MLE) is used to derive the parameters of the HVS model. However, MLE makes a number of assumptions and unfortunately some of these assumptions do not hold. Discriminative training, without making such assumptions, can improve the performance of the HVS model by discriminating the correct hypothesis from the competing hypotheses. Experiments have been conducted in two domains: the travel domain for the semantic parsing task using the DARPA Communicator data and the Air Travel Information Services (ATIS) data and the bioinformatics domain for the information extraction task using the GENIA corpus. The results demonstrate modest improvements of the performance of the HVS model using discriminative training. In the travel domain, discriminative training of the HVS model gives a relative error reduction rate of 31 percent in F-measure when compared with MLE on the DARPA Communicator data and 9 percent on the ATIS data. In the bioinformatics domain, a relative error reduction rate of 4 percent in F-measure is achieved on the GENIA corpus.

A hybrid generative/discriminative framework to train a semantic parser from an un-annotated corpus

We propose a hybrid generative/discriminative framework for semantic parsing which combines the hidden vector state (HVS) model and the hidden Markov support vector machines (HM-SVMs). The HVS model is an extension of the basic discrete Markov model in which context is encoded as a stack-oriented state vector. The HM-SVMs combine the advantages of the hidden Markov models and the support vector machines. By employing a modified K-means clustering method, a small set of most representative sentences can be automatically selected from an un-annotated corpus. These sentences together with their abstract annotations are used to train an HVS model which could be subsequently applied on the whole corpus to generate semantic parsing results. The most confident semantic parsing results are selected to generate a fully-annotated corpus which is used to train the HM-SVMs. The proposed framework has been tested on the DARPA Communicator Data. Experimental results show that an improvement over the baseline HVS parser has been observed using the hybrid framework. When compared with the HM-SVMs trained from the fully-annotated corpus, the hybrid framework gave a comparable performance with only a small set of lightly annotated sentences. © 2008. Licensed under the Creative Commons.

Feature LDA:a supervised topic model for automatic detection of web API documentations from the web

Web APIs have gained increasing popularity in recent Web service technology development owing to its simplicity of technology stack and the proliferation of mashups. However, efficiently discovering Web APIs and the relevant documentations on the Web is still a challenging task even with the best resources available on the Web. In this paper we cast the problem of detecting the Web API documentations as a text classification problem of classifying a given Web page as Web API associated or not. We propose a supervised generative topic model called feature latent Dirichlet allocation (feaLDA) which offers a generic probabilistic framework for automatic detection of Web APIs. feaLDA not only captures the correspondence between data and the associated class labels, but also provides a mechanism for incorporating side information such as labelled features automatically learned from data that can effectively help improving classification performance. Extensive experiments on our Web APIs documentation dataset shows that the feaLDA model outperforms three strong supervised baselines including naive Bayes, support vector machines, and the maximum entropy model, by over 3% in classification accuracy. In addition, feaLDA also gives superior performance when compared against other existing supervised topic models.

Bragg grating fast tunable filter for wavelength division multiplexing

A Bragg grating fast tunable filter prototype working over a linear tuning range of 45 nm with a maximum tuning speed of 21 nm/ms has been realized. The tunable filter system is based on two piezoelectric stack actuators moving a mechanical device thus compressing an apodized fiber Bragg grating. The filter allows both traction and compression and can work in transmission and in reflection. It is designed to work with a channel spacing of 100 GHz according to the ITU specifications for wavelength division multiplexing systems.

Ontologies for crisis management:a review of state of the art in ontology design and usability

The growing use of a variety of information systems in crisis management both by non-governmental organizations (NGOs) and emergency management agencies makes the challenges of information sharing and interoperability increasingly important. The use of semantic web technologies is a growing area and is a technology stack specifically suited to these challenges. This paper presents a review of ontologies, vocabularies and taxonomies that are useful in crisis management systems. We identify the different subject areas relevant to crisis management based on a review of the literature. The different ontologies and vocabularies available are analysed in terms of their coverage, design and usability. We also consider the use cases for which they were designed and the degree to which they follow a variety of standards. While providing comprehensive ontologies for the crisis domain is not feasible or desirable there is considerable scope to develop ontologies for the subject areas not currently covered and for the purposes of interoperability.

Investigating the structure of biomass-derived non-graphitizing mesoporous carbons by electron energy loss spectroscopy in the transmission electron microscope and X-ray photoelectron spectroscopy

We have investigated the microstructure and bonding of two biomass-based porous carbon chromatographic stationary phase materials (alginic acid-derived Starbon® and calcium alginate-derived mesoporous carbon spheres (AMCS) and a commercial porous graphitic carbon (PGC), using high resolution transmission electron microscopy, electron energy loss spectroscopy (EELS), N2 porosimetry and X-ray photoelectron spectroscopy (XPS). The planar carbon sp -content of all three material types is similar to that of traditional nongraphitizing carbon although, both biomass-based carbon types contain a greater percentage of fullerene character (i.e. curved graphene sheets) than a non-graphitizing carbon pyrolyzed at the same temperature. This is thought to arise during the pyrolytic breakdown of hexauronic acid residues into C5 intermediates. Energy dispersive X-ray and XPS analysis reveals a homogeneous distribution of calcium in the AMCS and a calcium catalysis mechanism is discussed. That both Starbon® and AMCS, with high-fullerene character, show chromatographic properties similar to those of a commercial PGC material with extended graphitic stacks, suggests that, for separations at the molecular level, curved fullerene- like and planar graphitic sheets are equivalent in PGC chromatography. In addition, variation in the number of graphitic layers suggests that stack depth has minimal effect on the retention mechanism in PGC chromatography. © 2013 Elsevier Ltd. All rights reserved.

Case Comment : Jones v Kernott:which road to Rome?

Jones v Kernott [2011] UKSC 53 (SC) Stack v Dowden [2007] UKHL 17; [2007] 2 A.C. 432 (HL). Casenote explores the implications of the ruling in Jones v Kernott by the Supreme Court and assesses its implications for English Property Law. Building on the commonwealth experience the casenote calls for the intorduction of a statutory scheme so as to allow courts to reallocate property rights to unmarried cohabitants in the event of relationship breakdown.

WLAN / WMAN Integration Architecture and Traffic Control for Voice Transmission

The popular technologies Wi-Fi and WiMAX for realization of WLAN and WMAN respectively are much different, but they could compliment each other providing competitive wireless access for voice traffic. The article develops the idea of WLAN/WMAN (Wi-Fi/WiMAX) integration. WiMAX is offering a backup for the traffic overflowing from Wi-Fi cells located into the WiMAX cell. Overflow process is improved by proposed rearrangement control algorithm applied to the Wi-Fi voice calls. There are also proposed analytical models for system throughput evaluation and verification of the effectiveness using WMAN as a backup for WLAN overflow traffic and the proposed call rearrangement algorithm as well.

AlGaInP red-emitting light emitting diode under extremely high pulsed pumping

Efficiency of commercial 620 nm InAlGaP Golden Dragon-cased high-power LEDs has been studied under extremely high pump current density up to 4.5 kA/cm2 and pulse duration from microsecond down to sub-nanosecond range. No efficiency decrease and negligible red shift of the emission wavelength is observed in the whole range of drive currents at nanosecond-range pulses with duty cycles well below 1%. Analysis of the pulse-duration dependence of the LED efficiency and emission spectrum suggests the active region overheating to be the major mechanism of the LED efficiency reduction at higher pumping, dominating over the electron overflow and Auger recombination.