The implementation of a design of experiments strategy to increase recombinant protein yields in yeast (review)

Biological processes are subject to the influence of numerous factors and their interactions, which may be non-linear in nature. In a recombinant protein production experiment, understanding the relative importance of these factors, and their influence on the yield and quality of the recombinant protein being produced, is an essential part of its optimisation. In many cases, implementing a design of experiments (DoE) approach has delivered this understanding. This chapter aims to provide the reader with useful pointers in applying a DoE strategy to improve the yields of recombinant yeast cultures.

Liposome formulation of poorly water soluble drugs:optimisation of drug loading and ESEM analysis of stability

Liposomes due to their biphasic characteristic and diversity in design, composition and construction, offer a dynamic and adaptable technology for enhancing drug solubility. Starting with equimolar egg-phosphatidylcholine (PC)/cholesterol liposomes, the influence of the liposomal composition and surface charge on the incorporation and retention of a model poorly water soluble drug, ibuprofen was investigated. Both the incorporation and the release of ibuprofen were influenced by the lipid composition of the multi-lamellar vesicles (MLV) with inclusion of the long alkyl chain lipid (dilignoceroyl phosphatidylcholine (C 24PC)) resulting in enhanced ibuprofen incorporation efficiency and retention. The cholesterol content of the liposome bilayer was also shown to influence ibuprofen incorporation with maximum ibuprofen incorporation efficiency achieved when 4 μmol of cholesterol was present in the MLV formulation. Addition of anionic lipid dicetylphosphate (DCP) reduced ibuprofen drug loading presumably due to electrostatic repulsive forces between the carboxyl group of ibuprofen and the anionic head-group of DCP. In contrast, the addition of 2 μmol of the cationic lipid stearylamine (SA) to the liposome formulation (PC:Chol - 16 μmol:4 μmol) increased ibuprofen incorporation efficiency by approximately 8%. However further increases of the SA content to 4 μmol and above reduced incorporation by almost 50% compared to liposome formulations excluding the cationic lipid. Environmental scanning electron microscopy (ESEM) was used to dynamically follow the changes in liposome morphology during dehydration to provide an alternative assay of liposome stability. ESEM analysis clearly demonstrated that ibuprofen incorporation improved the stability of PC:Chol liposomes as evidenced by an increased resistance to coalescence during dehydration. These finding suggest a positive interaction between amphiphilic ibuprofen molecules and the bilayer structure of the liposome. © 2004 Elsevier B.V. All rights reserved.

Requirement for and optimisation of premium intraocular lenses

Premium intraocular lenses (IOLs) aim to surgically correct astigmatism and presbyopia following cataract extraction, optimising vision and eliminating the need for cataract surgery in later years. It is usual to fully correct astigmatism and to provide visual correction for distance and near when prescribing spectacles and contact lenses, however for correction with the lens implanted during cataract surgery, patients are required to purchase the premium IOLs and pay surgery fees outside the National Health Service in the UK. The benefit of using toric IOLs was thus demonstrated, both in standard visual tests and real-world situations. Orientation of toric IOLs during implantation is critical and the benefit of using conjunctival blood vessels for alignment was shown. The issue of centration of IOLs relative to the pupil was also investigated, showing changes with the amount of dilation and repeat dilation evaluation, which must be considered during surgery to optimize the visual performance of premium IOLs. Presbyopia is a global issue, of growing importance as life expectancy increases, with no real long-term cure. Despite enhanced lifestyles, changes in diet and improved medical care, presbyopia still presents in modern life as a significant visual impairment. The onset of presbyopia was found to vary with risk factors including alcohol consumption, smoking, UV exposure and even weight as well as age. A new technique to make measurement of accommodation more objective and robust was explored, although needs for further design modifications were identified. Due to dysphotopsia and lack of intermediate vision through most multifocal IOL designs, the development of a trifocal IOL was shown to minimize these aspects. The current thesis, therefore, emphasises the challenges of premium IOL surgery and need for refinement for optimum visual outcome in addition to outlining how premium IOLs may provide long-term and successful correction of astigmatism and presbyopia.

Supply chain optimisation of pyrolysis plant deployment using goal programming

This paper presents a goal programming model to optimise the deployment of pyrolysis plants in Punjab, India. Punjab has an abundance of waste straw and pyrolysis can convert this waste into alternative bio-fuels, which will facilitate the provision of valuable energy services and reduce open field burning. A goal programming model is outlined and demonstrated in two case study applications: small scale operations in villages and large scale deployment across Punjab's districts. To design the supply chain, optimal decisions for location, size and number of plants, downstream energy applications and feedstocks processed are simultaneously made based on stakeholder requirements for capital cost, payback period and production cost of bio-oil and electricity. The model comprises quantitative data obtained from primary research and qualitative data gathered from farmers and potential investors. The Punjab district of Fatehgarh Sahib is found to be the ideal location to initially utilise pyrolysis technology. We conclude that goal programming is an improved method over more conventional methods used in the literature for project planning in the field of bio-energy. The model and findings developed from this study will be particularly valuable to investors, plant developers and municipalities interested in waste to energy in India and elsewhere. © 2014 Elsevier Ltd. All rights reserved.

Design and fabrication of micro-structured waveguides in lithium niobate

We review our recent work on the numerical design and optimisation of buried, micro-structured waveguides (WGs) that can be formed in a lithium niobate (LiNbO3) crystal by the method of direct femtosecond laser inscription. We also report on the possibility of fabricating such WGs using a high-repetition-rate, chirped-pulse oscillator system. Refractive index contrasts as high as -0.0127 have been achieved for individual modification tracks. The results pave the way for developing micro-structured WGs with low-loss operation across a wide spectral range, extending into the mid-infrared region up to the end of the transparency range of the host material. © 2014 IEEE.

The implementation of a design of experiments strategy to increase recombinant protein yields in yeast (review)

Biological processes are subject to the influence of numerous factors and their interactions, which may be non-linear in nature. In a recombinant protein production experiment, understanding the relative importance of these factors, and their influence on the yield and quality of the recombinant protein being produced, is an essential part of its optimisation. In many cases, implementing a design of experiments (DoE) approach has delivered this understanding. This chapter aims to provide the reader with useful pointers in applying a DoE strategy to improve the yields of recombinant yeast cultures.

Exposing market mechanism design trade-offs via multi-objective evolutionary search

Market mechanisms are a means by which resources in contention can be allocated between contending parties, both in human economies and those populated by software agents. Designing such mechanisms has traditionally been carried out by hand, and more recently by automation. Assessing these mechanisms typically involves them being evaluated with respect to multiple conflicting objectives, which can often be nonlinear, noisy, and expensive to compute. For typical performance objectives, it is known that designed mechanisms often fall short on being optimal across all objectives simultaneously. However, in all previous automated approaches, either only a single objective is considered, or else the multiple performance objectives are combined into a single objective. In this paper we do not aggregate objectives, instead considering a direct, novel application of multi-objective evolutionary algorithms (MOEAs) to the problem of automated mechanism design. This allows the automatic discovery of trade-offs that such objectives impose on mechanisms. We pose the problem of mechanism design, specifically for the class of linear redistribution mechanisms, as a naturally existing multi-objective optimisation problem. We apply a modified version of NSGA-II in order to design mechanisms within this class, given economically relevant objectives such as welfare and fairness. This application of NSGA-II exposes tradeoffs between objectives, revealing relationships between them that were otherwise unknown for this mechanism class. The understanding of the trade-off gained from the application of MOEAs can thus help practitioners with an insightful application of discovered mechanisms in their respective real/artificial markets.

Achievable performance gain of IEEE 802.11 multi-rate link adaptation algorithm with cross-layer design

Link quality-based rate adaptation has been widely used for IEEE 802.11 networks. However, network performance is affected by both link quality and random channel access. Selection of transmit modes for optimal link throughput can cause medium access control (MAC) throughput loss. In this paper, we investigate this issue and propose a generalised cross-layer rate adaptation algorithm. It considers jointly link quality and channel access to optimise network throughput. The objective is to examine the potential benefits by cross-layer design. An efficient analytic model is proposed to evaluate rate adaptation algorithms under dynamic channel and multi-user access environments. The proposed algorithm is compared to link throughput optimisation-based algorithm. It is found rate adaptation by optimising link layer throughput can result in large performance loss, which cannot be compensated by the means of optimising MAC access mechanism alone. Results show cross-layer design can achieve consistent and considerable performance gains of up to 20%. It deserves to be exploited in practical design for IEEE 802.11 networks.

Integrated optimisation of surface roughness and tool performance when face milling 416 SS

Tool life is an important factor to be considered during the optimisation of a machining process since cutting parameters can be adjusted to optimise tool changing, reducing cost and time of production. Also the performance of a tool is directly linked to the generated surface roughness and this is important in cases where there are strict surface quality requirements. The prediction of tool life and the resulting surface roughness in milling operations has attracted considerable research efforts. The research reported herein is focused on defining the influence of milling cutting parameters such as cutting speed, feed rate and axial depth of cut, on three major tool performance parameters namely, tool life, material removal and surface roughness. The research is seeking to define methods that will allow the selection of optimal parameters for best tool performance when face milling 416 stainless steel bars. For this study the Taguchi method was applied in a special design of an orthogonal array that allows studying the entire parameter space with only a number of experiments representing savings in cost and time of experiments. The findings were that the cutting speed has the most influence on tool life and surface roughness and very limited influence on material removal. By last tool life can be judged either from tool life or volume of material removal.

Powertrain optimisation in a hybrid electric bus

This paper describes the use of a formal optimisation procedure to optimise a plug-in hybrid electric bus using two different case studies to meet two different performance criteria; minimum journey cost and maximum battery life. The approach is to choose a commercially available vehicle and seek to improve its performance by varying key design parameters. Central to this approach is the ability to develop a representative backward facing model of the vehicle in MATLAB/Simulink along with appropriate optimisation objective and penalty functions. The penalty functions being the margin by which a particular design fails to meet the performance specification. The model is validated against data collected from an actual vehicle and is used to estimate the vehicle performance parameters in a model-in-the-loop process within an optimisation routine. For the purposes of this paper, the journey cost/battery life over a drive cycle is optimised whilst other performance indices are met (or exceeded). Among the available optimisation methods, Powell's method and Simulated Annealing are adopted. The results show this method as a valid alternative modelling approach to vehicle powertrain optimisation. © 2012 IEEE.

An application of evolutionary algorithms for WAG optimisation in the Norne Field

Water-alternating-gas (WAG) is an enhanced oil recovery method combining the improved macroscopic sweep of water flooding with the improved microscopic displacement of gas injection. The optimal design of the WAG parameters is usually based on numerical reservoir simulation via trial and error, limited by the reservoir engineer’s availability. Employing optimisation techniques can guide the simulation runs and reduce the number of function evaluations. In this study, robust evolutionary algorithms are utilized to optimise hydrocarbon WAG performance in the E-segment of the Norne field. The first objective function is selected to be the net present value (NPV) and two global semi-random search strategies, a genetic algorithm (GA) and particle swarm optimisation (PSO) are tested on different case studies with different numbers of controlling variables which are sampled from the set of water and gas injection rates, bottom-hole pressures of the oil production wells, cycle ratio, cycle time, the composition of the injected hydrocarbon gas (miscible/immiscible WAG) and the total WAG period. In progressive experiments, the number of decision-making variables is increased, increasing the problem complexity while potentially improving the efficacy of the WAG process. The second objective function is selected to be the incremental recovery factor (IRF) within a fixed total WAG simulation time and it is optimised using the same optimisation algorithms. The results from the two optimisation techniques are analyzed and their performance, convergence speed and the quality of the optimal solutions found by the algorithms in multiple trials are compared for each experiment. The distinctions between the optimal WAG parameters resulting from NPV and oil recovery optimisation are also examined. This is the first known work optimising over this complete set of WAG variables. The first use of PSO to optimise a WAG project at the field scale is also illustrated. Compared to the reference cases, the best overall values of the objective functions found by GA and PSO were 13.8% and 14.2% higher, respectively, if NPV is optimised over all the above variables, and 14.2% and 16.2% higher, respectively, if IRF is optimised.

Simultaneous optimisation of structural topology and material grading using level set method

Peer reviewed

Optimisation of a ground anchor head for NDT monitoring

Peer reviewed

Improving photofermentative hydrogen production through metabolic engineering and DOE (Design of Experiments)

A l’heure actuelle, les biocarburants renouvelables et qui ne nuit pas à l'environnement sont à l'étude intensive en raison de l'augmentation des problèmes de santé et de la diminution des combustibles fossiles. H2 est l'un des candidats les plus prometteurs en raison de ses caractéristiques uniques, telles que la densité d'énergie élevée et la génération faible ou inexistante de polluants. Une façon attrayante pour produire la H2 est par les bactéries photosynthétiques qui peuvent capter l'énergie lumineuse pour actionner la production H2 avec leur système de nitrogénase. L'objectif principal de cette étude était d'améliorer le rendement de H2 des bactéries photosynthétiques pourpres non sulfureuses utilisant une combinaison de génie métabolique et le plan des expériences. Une hypothèse est que le rendement en H2 pourrait être améliorée par la redirection de flux de cycle du Calvin-Benson-Bassham envers du système de nitrogénase qui catalyse la réduction des protons en H2. Ainsi, un PRK, phosphoribulose kinase, mutant « knock-out » de Rhodobacter capsulatus JP91 a été créé. L’analyse de la croissance sur des différentes sources de carbone a montré que ce mutant ne peut croître qu’avec l’acétate, sans toutefois produire d' H2. Un mutant spontané, YL1, a été récupéré qui a retenu l'cbbP (codant pour PRK) mutation d'origine, mais qui avait acquis la capacité de se développer sur le glucose et produire H2. Une étude de la production H2 sous différents niveaux d'éclairage a montré que le rendement d’YL1 était de 20-40% supérieure à la souche type sauvage JP91. Cependant, il n'y avait pas d'amélioration notable du taux de production de H2. Une étude cinétique a montré que la croissance et la production d'hydrogène sont fortement liées avec des électrons à partir du glucose principalement dirigés vers la production de H2 et la formation de la biomasse. Sous des intensités lumineuses faibles à intermédiaires, la production d'acides organiques est importante, ce qui suggère une nouvelle amélioration additionnel du rendement H2 pourrait être possible grâce à l'optimisation des processus. Dans une série d'expériences associées, un autre mutant spontané, YL2, qui a un phénotype similaire à YL1, a été testé pour la croissance dans un milieu contenant de l'ammonium. Les résultats ont montré que YL2 ne peut croître que avec de l'acétate comme source de carbone, encore une fois, sans produire de H2. Une incubation prolongée dans les milieux qui ne supportent pas la croissance de YL2 a permis l'isolement de deux mutants spontanés secondaires intéressants, YL3 et YL4. L'analyse par empreint du pied Western a montré que les deux souches ont, dans une gamme de concentrations d'ammonium, l'expression constitutive de la nitrogénase. Les génomes d’YL2, YL3 et YL4 ont été séquencés afin de trouver les mutations responsables de ce phénomène. Fait intéressant, les mutations de nifA1 et nifA2 ont été trouvés dans les deux YL3 et YL4. Il est probable qu'un changement conformationnel de NifA modifie l'interaction protéine-protéine entre NifA et PII protéines (telles que GlnB ou GlnK), lui permettant d'échapper à la régulation par l'ammonium, et donc d'être capable d'activer la transcription de la nitrogénase en présence d'ammonium. On ignore comment le nitrogénase synthétisé est capable de maintenir son activité parce qu’en théorie, il devrait également être soumis à une régulation post-traductionnelle par ammonium. Une autre preuve pourrait être obtenue par l'étude du transcriptome d’YL3 et YL4. Une première étude sur la production d’ H2 par YL3 et YL4 ont montré qu'ils sont capables d’une beaucoup plus grande production d'hydrogène que JP91 en milieu d'ammonium, qui ouvre la porte pour les études futures avec ces souches en utilisant des déchets contenant de l'ammonium en tant que substrats. Enfin, le reformage biologique de l'éthanol à H2 avec la bactérie photosynthétique, Rhodopseudomonas palustris CGA009 a été examiné. La production d'éthanol avec fermentation utilisant des ressources renouvelables microbiennes a été traitée comme une technique mature. Cependant, la plupart des études du reformage de l'éthanol à H2 se sont concentrés sur le reformage chimique à la vapeur, ce qui nécessite généralement une haute charge énergetique et résultats dans les émissions de gaz toxiques. Ainsi le reformage biologique de l'éthanol à H2 avec des bactéries photosynthétiques, qui peuvent capturer la lumière pour répondre aux besoins énergétiques de cette réaction, semble d’être plus prometteuse. Une étude précédente a démontré la production d'hydrogène à partir d'éthanol, toutefois, le rendement ou la durée de cette réaction n'a pas été examiné. Une analyse RSM (méthode de surface de réponse) a été réalisée dans laquelle les concentrations de trois facteurs principaux, l'intensité lumineuse, de l'éthanol et du glutamate ont été variés. Nos résultats ont montré que près de 2 moles de H2 peuvent être obtenus à partir d'une mole d'éthanol, 33% de ce qui est théoriquement possible.

Conception, caractérisation et optimisation de SPAD en technologie Dalsa HV CMOS 0.8 μm pour intégration dans un 3D-SiPM

Résumé : Les photodiodes à avalanche monophotonique (SPAD) sont d'intérêts pour les applications requérant la détection de photons uniques avec une grande résolution temporelle, comme en physique des hautes énergies et en imagerie médicale. En fait, les matrices de SPAD, souvent appelés photomultiplicateurs sur silicium (SiPM), remplacent graduellement les tubes photomultiplicateurs (PMT) et les photodiodes à avalanche (APD). De plus, il y a une tendance à utiliser les matrices de SPAD en technologie CMOS afin d'obtenir des pixels intelligents optimisés pour la résolution temporelle. La fabrication de SPAD en technologie CMOS commerciale apporte plusieurs avantages par rapport aux procédés optoélectroniques comme le faible coût, la capacité de production, l'intégration d'électronique et la miniaturisation des systèmes. Cependant, le défaut principal du CMOS est le manque de flexibilité de conception au niveau de l'architecture du SPAD, causé par le caractère fixe et standardisé des étapes de fabrication en technologie CMOS. Un autre inconvénient des matrices de SPAD CMOS est la perte de surface photosensible amenée par la présence de circuits CMOS. Ce document présente la conception, la caractérisation et l'optimisation de SPAD fabriqués dans une technologie CMOS commerciale (Teledyne DALSA 0.8µm HV CMOS - TDSI CMOSP8G). Des modifications de procédé sur mesure ont été introduites en collaboration avec l'entreprise CMOS pour optimiser les SPAD tout en gardant la compatibilité CMOS. Les matrices de SPAD produites sont dédiées à être intégrées en 3D avec de l'électronique CMOS économique (TDSI) ou avec de l'électronique CMOS submicronique avancée, produisant ainsi un SiPM 3D numérique. Ce SiPM 3D innovateur vise à remplacer les PMT, les APD et les SiPM commerciaux dans les applications à haute résolution temporelle. L'objectif principal du groupe de recherche est de développer un SiPM 3D avec une résolution temporelle de 10 ps pour usage en physique des hautes énergies et en imagerie médicale. Ces applications demandent des procédés fiables avec une capacité de production certifiée, ce qui justifie la volonté de produire le SiPM 3D avec des technologies CMOS commerciales. Ce mémoire étudie la conception, la caractérisation et l'optimisation de SPAD fabriqués en technologie TDSI-CMOSP8G.