High-throughput manufacturing of size-tuned liposomes by a new microfluidics method using enhanced statistical tools for characterization

Microfluidics has recently emerged as a new method of manufacturing liposomes, which allows for reproducible mixing in miliseconds on the nanoliter scale. Here we investigate microfluidics-based manufacturing of liposomes. The aim of these studies was to assess the parameters in a microfluidic process by varying the total flow rate (TFR) and the flow rate ratio (FRR) of the solvent and aqueous phases. Design of experiment and multivariate data analysis were used for increased process understanding and development of predictive and correlative models. High FRR lead to the bottom-up synthesis of liposomes, with a strong correlation with vesicle size, demonstrating the ability to in-process control liposomes size; the resulting liposome size correlated with the FRR in the microfluidics process, with liposomes of 50 nm being reproducibly manufactured. Furthermore, we demonstrate the potential of a high throughput manufacturing of liposomes using microfluidics with a four-fold increase in the volumetric flow rate, maintaining liposome characteristics. The efficacy of these liposomes was demonstrated in transfection studies and was modelled using predictive modeling. Mathematical modelling identified FRR as the key variable in the microfluidic process, with the highest impact on liposome size, polydispersity and transfection efficiency. This study demonstrates microfluidics as a robust and high-throughput method for the scalable and highly reproducible manufacture of size-controlled liposomes. Furthermore, the application of statistically based process control increases understanding and allows for the generation of a design-space for controlled particle characteristics.

Control of the properties of micro-structured waveguides in LiNbO3 fabricated by direct femtosecond laser inscription

This paper reports on buried waveguides fabricated in lithium niobate (LN) by the method of direct femtosecond (fs) laser inscription. 5% MgO doped LiNbO3 was chosen as the host material because of its high quality and damage threshold, as well as relatively low cost. Direct fs inscription by astigmatically shaped beam in crystals usually produces multiple 'smooth' tracks (with reduced refractive index), which encircle the light guiding 'core', thus creating a depressed cladding WG. A high-repetition rate fs laser system was used for inscription at a depth of approximately 500 μm. Using numerical modelling, it was demonstrated that the properties of fs-written WGs can be controlled by the WG geometry. Buried, depressed-cladding WGs in LN host with circular cross-section were also demonstrated. Combining control over the WG dispersion with quasi-phase matching will allow various ultralow-pump-power, highly-efficient, nonlinear light-guiding devices - all in an integrated optics format.

Neural Network Based Optimal Control with Constraints

In the present paper the problems of the optimal control of systems when constraints are imposed on the control is considered. The optimality conditions are given in the form of Pontryagin’s maximum principle. The obtained piecewise linear function is approximated by using feedforward neural network. A numerical example is given.

Optimal Control of a Second Order Parabolic Heat Equation

In this paper, we are concerned with the optimal control boundary control of a second order parabolic heat equation. Using the results in [Evtushenko, 1997] and spatial central finite difference with diagonally implicit Runge-Kutta method (DIRK) is applied to solve the parabolic heat equation. The conjugate gradient method (CGM) is applied to solve the distributed control problem. Numerical results are reported.

Robot Control Using Inductive, Deductive and Case Based Reasoning

The paper deals with a problem of intelligent system’s design for complex environments. There is discussed a possibility to integrate several technologies into one basic structure that could form a kernel of an autonomous intelligent robotic system. One alternative structure is proposed in order to form a basis of an intelligent system that would be able to operate in complex environments. The proposed structure is very flexible because of features that allow adapting via learning and adjustment of the used knowledge. Therefore, the proposed structure may be used in environments with stochastic features such as hardly predictable events or elements. The basic elements of the proposed structure have found their implementation in software system and experimental robotic system. The software system as well as the robotic system has been used for experimentation in order to validate the proposed structure - its functionality, flexibility and reliability. Both of them are presented in the paper. The basic features of each system are presented as well. The most important results of experiments are outlined and discussed at the end of the paper. Some possible directions of further research are also sketched at the end of the paper.

Adaptive Control and Multi-agent Interface for Infotelecommunication Systems of New Generation

Problems for intellectualisation for man-machine interface and methods of self-organization for network control in multi-agent infotelecommunication systems have been discussed. Architecture and principles for construction of network and neural agents for telecommunication systems of new generation have been suggested. Methods for adaptive and multi-agent routing for information flows by requests of external agents- users of global telecommunication systems and computer networks have been described.

Editorial overview:new protein production tools for structural biology

Full text: The idea of producing proteins from recombinant DNA hatched almost half a century ago. In his PhD thesis, Peter Lobban foresaw the prospect of inserting foreign DNA (from any source, including mammalian cells) into the genome of a λ phage in order to detect and recover protein products from Escherichia coli [ 1 and 2]. Only a few years later, in 1977, Herbert Boyer and his colleagues succeeded in the first ever expression of a peptide-coding gene in E. coli — they produced recombinant somatostatin [ 3] followed shortly after by human insulin. The field has advanced enormously since those early days and today recombinant proteins have become indispensable in advancing research and development in all fields of the life sciences. Structural biology, in particular, has benefitted tremendously from recombinant protein biotechnology, and an overwhelming proportion of the entries in the Protein Data Bank (PDB) are based on heterologously expressed proteins. Nonetheless, synthesizing, purifying and stabilizing recombinant proteins can still be thoroughly challenging. For example, the soluble proteome is organized to a large part into multicomponent complexes (in humans often comprising ten or more subunits), posing critical challenges for recombinant production. A third of all proteins in cells are located in the membrane, and pose special challenges that require a more bespoke approach. Recent advances may now mean that even these most recalcitrant of proteins could become tenable structural biology targets on a more routine basis. In this special issue, we examine progress in key areas that suggests this is indeed the case. Our first contribution examines the importance of understanding quality control in the host cell during recombinant protein production, and pays particular attention to the synthesis of recombinant membrane proteins. A major challenge faced by any host cell factory is the balance it must strike between its own requirements for growth and the fact that its cellular machinery has essentially been hijacked by an expression construct. In this context, Bill and von der Haar examine emerging insights into the role of the dependent pathways of translation and protein folding in defining high-yielding recombinant membrane protein production experiments for the common prokaryotic and eukaryotic expression hosts. Rather than acting as isolated entities, many membrane proteins form complexes to carry out their functions. To understand their biological mechanisms, it is essential to study the molecular structure of the intact membrane protein assemblies. Recombinant production of membrane protein complexes is still a formidable, at times insurmountable, challenge. In these cases, extraction from natural sources is the only option to prepare samples for structural and functional studies. Zorman and co-workers, in our second contribution, provide an overview of recent advances in the production of multi-subunit membrane protein complexes and highlight recent achievements in membrane protein structural research brought about by state-of-the-art near-atomic resolution cryo-electron microscopy techniques. E. coli has been the dominant host cell for recombinant protein production. Nonetheless, eukaryotic expression systems, including yeasts, insect cells and mammalian cells, are increasingly gaining prominence in the field. The yeast species Pichia pastoris, is a well-established recombinant expression system for a number of applications, including the production of a range of different membrane proteins. Byrne reviews high-resolution structures that have been determined using this methylotroph as an expression host. Although it is not yet clear why P. pastoris is suited to producing such a wide range of membrane proteins, its ease of use and the availability of diverse tools that can be readily implemented in standard bioscience laboratories mean that it is likely to become an increasingly popular option in structural biology pipelines. The contribution by Columbus concludes the membrane protein section of this volume. In her overview of post-expression strategies, Columbus surveys the four most common biochemical approaches for the structural investigation of membrane proteins. Limited proteolysis has successfully aided structure determination of membrane proteins in many cases. Deglycosylation of membrane proteins following production and purification analysis has also facilitated membrane protein structure analysis. Moreover, chemical modifications, such as lysine methylation and cysteine alkylation, have proven their worth to facilitate crystallization of membrane proteins, as well as NMR investigations of membrane protein conformational sampling. Together these approaches have greatly facilitated the structure determination of more than 40 membrane proteins to date. It may be an advantage to produce a target protein in mammalian cells, especially if authentic post-translational modifications such as glycosylation are required for proper activity. Chinese Hamster Ovary (CHO) cells and Human Embryonic Kidney (HEK) 293 cell lines have emerged as excellent hosts for heterologous production. The generation of stable cell-lines is often an aspiration for synthesizing proteins expressed in mammalian cells, in particular if high volumetric yields are to be achieved. In his report, Buessow surveys recent structures of proteins produced using stable mammalian cells and summarizes both well-established and novel approaches to facilitate stable cell-line generation for structural biology applications. The ambition of many biologists is to observe a protein's structure in the native environment of the cell itself. Until recently, this seemed to be more of a dream than a reality. Advances in nuclear magnetic resonance (NMR) spectroscopy techniques, however, have now made possible the observation of mechanistic events at the molecular level of protein structure. Smith and colleagues, in an exciting contribution, review emerging ‘in-cell NMR’ techniques that demonstrate the potential to monitor biological activities by NMR in real time in native physiological environments. A current drawback of NMR as a structure determination tool derives from size limitations of the molecule under investigation and the structures of large proteins and their complexes are therefore typically intractable by NMR. A solution to this challenge is the use of selective isotope labeling of the target protein, which results in a marked reduction of the complexity of NMR spectra and allows dynamic processes even in very large proteins and even ribosomes to be investigated. Kerfah and co-workers introduce methyl-specific isotopic labeling as a molecular tool-box, and review its applications to the solution NMR analysis of large proteins. Tyagi and Lemke next examine single-molecule FRET and crosslinking following the co-translational incorporation of non-canonical amino acids (ncAAs); the goal here is to move beyond static snap-shots of proteins and their complexes and to observe them as dynamic entities. The encoding of ncAAs through codon-suppression technology allows biomolecules to be investigated with diverse structural biology methods. In their article, Tyagi and Lemke discuss these approaches and speculate on the design of improved host organisms for ‘integrative structural biology research’. Our volume concludes with two contributions that resolve particular bottlenecks in the protein structure determination pipeline. The contribution by Crepin and co-workers introduces the concept of polyproteins in contemporary structural biology. Polyproteins are widespread in nature. They represent long polypeptide chains in which individual smaller proteins with different biological function are covalently linked together. Highly specific proteases then tailor the polyprotein into its constituent proteins. Many viruses use polyproteins as a means of organizing their proteome. The concept of polyproteins has now been exploited successfully to produce hitherto inaccessible recombinant protein complexes. For instance, by means of a self-processing synthetic polyprotein, the influenza polymerase, a high-value drug target that had remained elusive for decades, has been produced, and its high-resolution structure determined. In the contribution by Desmyter and co-workers, a further, often imposing, bottleneck in high-resolution protein structure determination is addressed: The requirement to form stable three-dimensional crystal lattices that diffract incident X-ray radiation to high resolution. Nanobodies have proven to be uniquely useful as crystallization chaperones, to coax challenging targets into suitable crystal lattices. Desmyter and co-workers review the generation of nanobodies by immunization, and highlight the application of this powerful technology to the crystallography of important protein specimens including G protein-coupled receptors (GPCRs). Recombinant protein production has come a long way since Peter Lobban's hypothesis in the late 1960s, with recombinant proteins now a dominant force in structural biology. The contributions in this volume showcase an impressive array of inventive approaches that are being developed and implemented, ever increasing the scope of recombinant technology to facilitate the determination of elusive protein structures. Powerful new methods from synthetic biology are further accelerating progress. Structure determination is now reaching into the living cell with the ultimate goal of observing functional molecular architectures in action in their native physiological environment. We anticipate that even the most challenging protein assemblies will be tackled by recombinant technology in the near future.

A Gradient-Type Optimization Technique for the Optimal Control for Schrodinger Equations

In this paper, we are considered with the optimal control of a schrodinger equation. Based on the formulation for the variation of the cost functional, a gradient-type optimization technique utilizing the finite difference method is then developed to solve the constrained optimization problem. Finally, a numerical example is given and the results show that the method of solution is robust.

Timed Transition Automata as Numerical Planning Domain

A general technique for transforming a timed finite state automaton into an equivalent automated planning domain based on a numerical parameter model is introduced. Timed transition automata have many applications in control systems and agents models; they are used to describe sequential processes, where actions are labelling by automaton transitions subject to temporal constraints. The language of timed words accepted by a timed automaton, the possible sequences of system or agent behaviour, can be described in term of an appropriate planning domain encapsulating the timed actions patterns and constraints. The time words recognition problem is then posed as a planning problem where the goal is to reach a final state by a sequence of actions, which corresponds to the timed symbols labeling the automaton transitions. The transformation is proved to be correct and complete and it is space/time linear on the automaton size. Experimental results shows that the performance of the planning domain obtained by transformation is scalable for real world applications. A major advantage of the planning based approach, beside of the solving the parsing problem, is to represent in a single automated reasoning framework problems of plan recognitions, plan synthesis and plan optimisation.

Semi-Smooth Newton Methods for the Time Optimal Control of Nonautonomous Ordinary Differential Equations

AMS Subj. Classification: 49J15, 49M15

Comparison between conventional and stochastic pinning control

Theoretical developments on pinning control of complex dynamical networks have mainly focused on the deterministic versions of the model dynamics. However, the dynamical behavior of most real networks is often affected by stochastic noise components. In this paper the pinning control of a stochastic version of the coupled map lattice network with spatiotemporal characteristics is studied. The control of these complex dynamical networks have functional uncertainty which should be considered when calculating stabilizing control signals. Two feedback control methods are considered: the conventional feedback control and modified stochastic feedback control. It is shown that the typically-used conventional control method suffers from the ignorance of model uncertainty leading to a reduction and potentially a collapse in the control efficiency. Numerical verification of the main result is provided for a chaotic coupled map lattice network. © 2011 IEEE.

Loss of Accuracy in Numerical Computations

Михаил М. Константинов, Петко Х. Петков - Разгледани са възможните катастрофални ефекти от неправилното използване на крайна машинна аритметика с плаваща точка. За съжаление, тази тема не винаги се разбира достатъчно добре от студентите по приложна и изчислителна математика, като положението в инженерните и икономическите специалности в никакъв случай не е по-добро. За преодоляване на този образователен пропуск тук сме разгледали главните виновници за загубата на точност при числените компютърни пресмятания. Надяваме се, че представените резултати ще помогнат на студентите и лекторите за по-добро разбиране и съответно за избягване на основните фактори, които могат да разрушат точността при компютърните числени пресмятания. Последното не е маловажно – числените катастрофи понякога стават истински, с големи щети и човешки жертви.

A Taxonomy of Big Data for Optimal Predictive Machine Learning and Data Mining

Big data comes in various ways, types, shapes, forms and sizes. Indeed, almost all areas of science, technology, medicine, public health, economics, business, linguistics and social science are bombarded by ever increasing flows of data begging to be analyzed efficiently and effectively. In this paper, we propose a rough idea of a possible taxonomy of big data, along with some of the most commonly used tools for handling each particular category of bigness. The dimensionality p of the input space and the sample size n are usually the main ingredients in the characterization of data bigness. The specific statistical machine learning technique used to handle a particular big data set will depend on which category it falls in within the bigness taxonomy. Large p small n data sets for instance require a different set of tools from the large n small p variety. Among other tools, we discuss Preprocessing, Standardization, Imputation, Projection, Regularization, Penalization, Compression, Reduction, Selection, Kernelization, Hybridization, Parallelization, Aggregation, Randomization, Replication, Sequentialization. Indeed, it is important to emphasize right away that the so-called no free lunch theorem applies here, in the sense that there is no universally superior method that outperforms all other methods on all categories of bigness. It is also important to stress the fact that simplicity in the sense of Ockham’s razor non-plurality principle of parsimony tends to reign supreme when it comes to massive data. We conclude with a comparison of the predictive performance of some of the most commonly used methods on a few data sets.

Identifying DNA-binding proteins by combining support vector machine and PSSM distance transformation

Background: DNA-binding proteins play a pivotal role in various intra- and extra-cellular activities ranging from DNA replication to gene expression control. Identification of DNA-binding proteins is one of the major challenges in the field of genome annotation. There have been several computational methods proposed in the literature to deal with the DNA-binding protein identification. However, most of them can't provide an invaluable knowledge base for our understanding of DNA-protein interactions. Results: We firstly presented a new protein sequence encoding method called PSSM Distance Transformation, and then constructed a DNA-binding protein identification method (SVM-PSSM-DT) by combining PSSM Distance Transformation with support vector machine (SVM). First, the PSSM profiles are generated by using the PSI-BLAST program to search the non-redundant (NR) database. Next, the PSSM profiles are transformed into uniform numeric representations appropriately by distance transformation scheme. Lastly, the resulting uniform numeric representations are inputted into a SVM classifier for prediction. Thus whether a sequence can bind to DNA or not can be determined. In benchmark test on 525 DNA-binding and 550 non DNA-binding proteins using jackknife validation, the present model achieved an ACC of 79.96%, MCC of 0.622 and AUC of 86.50%. This performance is considerably better than most of the existing state-of-the-art predictive methods. When tested on a recently constructed independent dataset PDB186, SVM-PSSM-DT also achieved the best performance with ACC of 80.00%, MCC of 0.647 and AUC of 87.40%, and outperformed some existing state-of-the-art methods. Conclusions: The experiment results demonstrate that PSSM Distance Transformation is an available protein sequence encoding method and SVM-PSSM-DT is a useful tool for identifying the DNA-binding proteins. A user-friendly web-server of SVM-PSSM-DT was constructed, which is freely accessible to the public at the web-site on http://bioinformatics.hitsz.edu.cn/PSSM-DT/.

Fiber bragg gratings:advances in fabrication process and tools

Successful commercialization of a technology such as Fiber Bragg Gratings requires the ability to manufacture devices repeatably, quickly and at low cost. Although the first report of photorefractive gratings was in 1978 it was not until 1993, when phase mask fabrication was demonstrated, that this became feasible. More recently, draw tower fabrication on a production level and grating writing through the polymer jacket have been realized; both important developments since they preserve the intrinsic strength of the fiber. Potentially the most significant recent development has been femtosecond laser inscription of gratings. Although not yet a commercial technology, it provides the means of writing multiple gratings in the optical core providing directional sensing capability in a single fiber. Femtosecond processing can also be used to machine the fiber to produce micronscale slots and holes enhancing the interaction between the light in the core and the surrounding medium. © 2011 Bentham Science Publishers Ltd. All rights reserved.