Construction and optimization of a CC49-based scFv-beta-lactamase fusion protein for ADEPT

CC49 is a clinically validated antibody with specificity for TAG-72, a carbohydrate epitope that is over-expressed and exposed on a large fraction of solid malignancies. We constructed a single chain fragment (scFv) based on CC49 and fused it to beta-lactamase. The first generation fusion protein, TAB2.4, was expressed at low levels in Escherichia coli and significant degradation was observed during production. We optimized the scFv domain of TAB2.4 by Combinatorial Consensus Mutagenesis (CCM). An improved variant TAB2.5 was identified that resulted in an almost 4-fold improved expression and 2.5 degrees higher thermostability relative to its parent molecule. Soluble TAB2.5 can be manufactured in low-density E.coli cultures at 120 mg/l. Our studies suggest that CCM is a rapid and efficient method to generate antibody fragments with improved stability and expression. The fusion protein TAB2.5 can be used for antibody directed enzyme prodrug therapy (ADEPT).     

Limitations of the terahertz photomixer

In the paper, Fourier transform has been used for calculations of a refractive index of dielectric samples measured in the terahertz photomixer arrangement. We considered measurement limitations caused by a sampling frequency and a photomixer bandwidth.     

Apparatus for the high temperature measurement of the Seebeck coefficient in thermoelectric materials

The Seebeck coefficient is a physical parameter routinely measured to identify the potential thermoelectric performance of a material. However, researchers employ a variety of techniques, conditions, and probe arrangements to measure the Seebeck coefficient, resulting in conflicting materials data. To compare and evaluate these methodologies, and to identify optimal Seebeck coefficient measurement protocols, we have developed an improved experimental apparatus to measure the Seebeck coefficient under multiple conditions and probe arrangements (300 K-1200 K). This paper will describe in detail the apparatus design and instrumentation, including a discussion of its capabilities and accuracy as measured through representative diagnostics. In addition, this paper will emphasize the techniques required to effectively manage uncertainty in high temperature Seebeck coefficient measurements.     

Application of lyophilization to prepare the nitrifying bacterial biofilm for imaging with scanning electron microscopy

Structure of bacterial biofilms may be investigated using several variants of scanning electron microscopy (SEM). We apply lyophilization to prepare nitrifying bacterial biofilm for conventional SEM imaging in high-vacuum mode (CSEM). We therefore replace standard biofilm fixation in glutaraldehyde cross-linking, ethanol dehydration, and critical-point drying (CPD) with less-invasive low-temperature drying by sublimation in vacuum. We compare this approach with: (1) standard preparation with glutaraldehyde fixation, ethanol dehydration, and CPD before CSEM, (2) cryo-sputter preparation of rapidly frozen biofilm in hydrated state (cryo-SEM), and (3) in situ observation without any sample pretreatment in environmental SEM. Combined imaging with these modalities revealed two distinct immobilization patterns on the polyurethane foam: (1) large irregular aggregates (flocs) of bacterial biofilm that exist as irregular biofilm fragments, rope-like structures, or biofilm layers on the foam surface; (2) biofilm threads adherent to the surface of polyurethane foam. Our results indicate that lyophilization was suitable for preservation of bacterial cells and many forms of structure of extracellular matrix. The lyophilized material could be imaged with high resolution (using CSEM) to generate structural information complementary to that obtained with other SEM techniques.     

Application of direct/inverse analysis to evaluate the structure integrity of concrete. Part I. Formulation for thermo-mechanical properties determination

Inverse analyses (IA) is used to model heat-flow characteristics in a continuum composed of two media. The process solves iteratively an optional number of design parameters. The solution also includes, (a) simulation of equipment and operator error due to data capture, and (b) sensitivity analysis of thermo-mechanical parameters.The analysis models test sample, which is sandwiched between a heating element and a reference material (of known thermal conductivity). This model is then validated by analysing actual thermographs recorded at the steady-state condition, the mean variation of which is subsequently used for IA.IA can then be applied in order to determine the best estimate of the thermo-mechanical properties of the sample in a particular controlled state of integrity or damage.     

Implementation of similarity model in the CSB system for chemical reaction predictions

A new algorithm for predicting the course of chemical reactions, implemented in the CSB system is presented. It allows to use of four various models for reaction generation, that lead to create different solution space according to the needs of the user. Depending on the chosen model one can expect solutions featuring great completeness or great reliability.