Aequorin-based functional assays for G-protein-coupled receptors,ion channels,and tyrosine kinase receptors

Aequorin is a photoprotein originating from jellyfish, whose luminescent activity is dependent on the concentration of calcium ions. Due to the high sensitivity and low background linked to luminescent assays, as well as to its absence of toxicity and its large linear dynamic range, aequorin has been used as an intracellular calcium indicator since its discovery in the early 1960s. The first applications of aequorin involved its microinjection in cells. The cloning of its gene in 1985 opened the way to the stable expression of aequorin in cell lines or even entire organisms. Here we present the validation of aequorin as a functional assay for the screening of G-protein-coupled receptors, ion channels, and tyrosine kinase receptors, as well as for their pharmacological characterization in agonist and antagonist detection assays. We optimized our cell suspension-based assay and determined that the most sensitive assay was performed at room temperature, with mitochondrially expressed aequorin and using coelenterazine derivative h for reconstitution of aequorin. The robustness of the assay and the current availability of luminometers with integrated injectors allow aequorin to fit perfectly with high throughput functional assays requirements.     

Configurations of NPD – Production Interfaces and Interface Integration Mechanisms

This paper describes and illustrates different configurations of the interface between new product development and production processes, including both intra–firm and inter–firm interfaces. These configurations are partly based on a process view of product innovation and partly on a structural view of product innovation. In addition to this typology of interfaces some integration mechanisms are described. The typology will serve as a basis for further research aimed at identifying consistent configurations of the different types of integration mechanisms that are available for industry to increase the effectiveness and efficiency of product innovation processes.     

Introduction by the conference editor

This paper deals with the problem of structuring the confusing pattern of problems of computerization. The approach to finding a relevant frame of reference for a discussion of the impact of computerization on qualities of work is to make some clear and basic distinctions by asking some very general questions and the look at the typical answers.The author's main argument is, that this approach leads to the following distinctions: Firstly, the distinction between three dimensions in the concept of work, which leads to the distinction between the structural, the functional, and the ethical aspects of computerization. Secondly, the distinction between three levels of society, which leads to the distinction between the societal, the social, and the psycho-physical aspects of computerization. Thirdly, the distinction between three types of questions regarding work, which leads to the distinction between the quantitative, the qualitative, and the utility aspects of computerization. However, these three times three times three aspects may be regarded by at least five different interest groups each asking at least four different interest groups each asking at least four different questions. No wonder that the problem is complex and the discussion may be confusing.The point is, that to find a reasonable pattern in this mess of problems we have to choose a human point of view and use a general framework of qualities of life for structuring the complex of problems. Essential to the human being are the relations between man and environment, and working environment should be regarded as a subsystem of environment in general. By doing that we find, that the change in the distribution of responsibility of doing the work between man and machine is the most remarkable impact of computerization.     

Isolation,characterization, and quantitative analysis of Microviridin J,a new Microcystis metabolite toxic to Daphnia

This paper describes the purification and characterization of microviridin J, a newly discovered metabolite of Microcystis that causes a lethal molting disruption in Daphnia spp., upon ingestion of living cyanobacterial cells. Microviridin J consists of an acetylated chain of 13 amino acids arranged in three rings and two side chains. Unlike other known isoforms of microviridin, microviridin J contains arginine that imparts a unique solution conformation characterized by proximal hydrophobic interactions between Arg and other regions of the molecule. This eventually results in the formation and stabilization of an additional ring system. Microviridin J potently inhibits porcine trypsin, bovine chymotrypsin, and daphnid trypsin-like proteases. The activity against trypsin is most likely due to Arg and its distinctive conformational interactions. Overall, the data presented for microviridin J emphasize once again the ability of cyanobacteria to produce numerous and potent environmental toxins.     

Determining the finite elasticity reference state from a loaded configuration

There are a number of situations where the deformed configuration of a body is known and it is necessary to determine the reference state. Previous methods developed to calculate the reference state involve the formulation of the finite elasticity equations in terms of the deformed configuration. This paper demonstrates that the undeformed reference state can be accurately determined from a deformed configuration and the associated loading conditions, by using conventional finite elasticity balance equations together with a solution procedure that treats the reference configuration as the unknowns. The mathematical theory behind the solution method is described, validated with an analytical solution, and verified using experimental studies on gel phantoms. The practical utility of this method is then demonstrated in the field of breast biomechanics. Copyright © 2007 John Wiley & Sons, Ltd.     

On non-linear unknown input observers–applied to lateral vehicle velocity estimation on banked roads

Unknown input observers (UIOs) are observers that have stable error dynamics that are independent of unknown inputs. This paper studies such observers for non-linear systems, and shows that the error dynamics for a non-linear UIO has the same structure as the error dynamics of a non-linear observer without unknown inputs. This result is first used to provide synthesis inequalities for UIOs for a class of non-linear systems, and secondly, to inspire the design of an observer for estimation of vehicle lateral velocity on banked roads.     

Design and analysis of gain-scheduled control using local controller networks

This paper describes a nonlinear control structure known as a local controller network. The structure consists of a weighted combination of a number of individual controllers, each of which is valid locally in the state space of the plant. Local controller designs are based upon local models valid in operating regimes which do not necessarily contain any physical equilibria. Consequently, the transient performance can be improved. Some 'scheduling' variables determine the current operating regime, and a validity function is assigned to each local controller. A 'feedforward' component may be used in each local controller in order to compensate directly for the operating-point-dependent model offsets. The application of the local controller network approach to a nonlinear control problem, that of longitudinal vehicle dynamics control, is described. A stability analysis for the discrete-time local controller network is given in this paper and the results are compared with known theoretical guidelines for related control approaches such as gain scheduling and feedback linearization.