Modification of human platelet adhesion on biomaterial surfaces by protein preadsorption under static and flow conditions

Biomaterial-induced thrombosis remains one of the main complications of vascular implant devices. Preadsorbed proteins on the biomaterial/blood interface will modify the adhesion and activation of platelets (PTLs) during the initial contact-phase. Our results clearly show that PTL-adherence on biomaterials is influenced not only by protein preadsorption, but also by flow conditions. The covalent coating of TCPS and glass by phosphorylcholine (PC) induces a significant decrease of PTL adhesion but leads to a slight, but nevertheless significant activation of PTL, which was detected by the induction of P-selectin expression using FACS analysis. Methodologically, the visualization of PTL adhesion gave more reliable results for measurement of PTL adhesion than the cell-enzyme immunoassay (EIA) for P-selectin. Human citrated plasma caused an inhibition of PTL. It is probable, that the contained sodium citrate may inhibit PTL adhesion by its calcium ion-binding capacity. The flow experiment as dynamic system is in our view absolutely essential for the evaluation of biomaterials for vascular prosthesis, and is in accordance with the international standards. The results of the experiments also suggest that investigations under static and flow conditions are needed to determine the influence of protein adsorption on mixed blood cell populations, for example, on PTL and PMN mixtures/co-cultures in order to achieve a better simulation of the in vivo situation.     

Polyplexes of polyethylenimine and per-N-methylated polyethylenimine-cytotoxicity and transfection efficiency

For non-viral gene delivery, the carriers for DNA transfer into cells must be vastly improved. The branched cationic polymer polyethylenimine has been described as an efficient gene carrier. However, polyethylenimine was demonstrated to mediate substantial cytotoxicity. Therefore, this study is aimed at investigating per-N-methylated polyethylenimine, which is thought to have a much lower cytotoxicity due to its lower charge density. Results from a gel retardation assay and laser light scattering indicated that per-N-methylated polyethylenimine condenses DNA into small and compact nanoparticles with a mean diameter <150 nm. Furthermore, polyplexes of polyethylenimine and per-N-methylated polyethylenimine with DNA had a positive zeta potential and the polymers protected DNA from nuclease-mediated digestion. The transfection efficiency of polyethylenimine and per-N-methylated polyethylenimine was tested in CHO-K1 cells. Using green fluorescent protein as reporter gene and flow cytometry analysis, we demonstrated that per-N-methylated polyethylenimine has a lower cytotoxicity, but also a significantly lower transfection efficiency. Using propidium iodide staining, we could additionally distinguish between viable and dead cells. At NP > or = 12, per-N-methylated polyethylenimine showed a much higher cell viability and the ratio of viable and transfected cells to dead and transfected cells was about 1.5 to 1.7 fold higher than for polyethylenimine. The results of cell viability from flow cytometry analysis were confirmed by the MTS assay. Using luciferase reporter gene for transfection experiments, the gene expression of per-N-methylated polyethylenimine was lower at NP 6, 12 and 18 as compared to polyethylenimine, but at NP 24 it yielded similar levels.     

In situ derivatization/solid-phase microextraction: determination of polar aromatic amines

A solid-phase microextraction GC/MS method for the trace determination of a wide variety of polar aromatic amines in aqueous samples was developed. Prior to extraction the analytes were derivatized directly in the aqueous solution by diazotation and subsequent iodination in a one-pot reaction. The derivatives were extracted by direct-SPME using a PDMS/DVB fiber and analyzed by GC/MS in the full-scan mode. By diazotation/iodination, the polarity of the analytes was significantly decreased and as a consequence extraction yields were dramatically improved. The derivatization proved to be suitable for strongly deactivated aromatic amines and even the very polar diamino compounds can efficiently be enriched after derivatization. We investigated 18 anilines comprising a wide range of functional groups, which could be determined simultaneously. The method was thoroughly validated, and the precision at a concentration of 0.5 microg/L was 3.8-11% relative standard deviation for nonnitrated analytes using aniline-d(5) as internal standard and 3.7-10% for nitroaromatic amines without internal standard. The in situ derivatization/SPME/GC/MS method was calibrated over the whole analytical procedure and was linear over 2 orders of magnitude. Using 10-mL samples, detection limits of 2-13 ng/L were achieved for 15 of the 18 analytes. For two aminodinitrotoluene isomers and a diaminonitrotoluene, detection limits ranged from 27 to 38 ng/L. By allowing quantification at the 0.1 microg/L level, analysis of all target compounds meets EU drinking water regulations. The method provides high sensitivity, robustness, and high sample throughput by automation. Finally, the method was applied to various real water samples and in wastewater from a former ammunition plant the contents of several aromatic amines were quantified.     

Large-Scale CFD Data Handling in a VR-Based Otorhinolaryngological CAS-System using a Linux-Cluster

Many of the operations to eliminate complaints concerning respiration impairments fail. In order to improve the success rate, it is important to recognize the responsiveness of the flow field within the nasal cavities. Therefore, we are developing a computer assisted surgery (CAS) system that combines computational fluid dynamics (CFD) and virtual reality (VR) technology. However, the primary prerequisite for VR-based applications is real-time interaction. A single graphics workstation is not capable of satisfying this condition and of simultaneously calculating flow features employing the huge CFD data set. In this paper, we will present our approach of a distributed system that relieves the load on the graphics workstation and makes use of an off-the-shelf parallel Linux cluster calculating streamlines. Moreover, we introduce first results and discuss remaining difficulties.     

An exact method to quantify the information transmitted by different mechanisms of correlational coding

We derive a new method to quantify the impact of correlated firing on the information transmitted by neuronal populations. This new method considers, in an exact way, the effects of high order spike train statistics, with no approximation involved, and it generalizes our previous work that was valid for short time windows and small populations. The new technique permits one to quantify the information transmitted if each cell were to convey fully independent information separately from the information available in the presence of synergy-redundancy effects. Synergy-redundancy effects are shown to arise from three possible contributions: a redundant contribution due to similarities in the mean response profiles of different cells; a synergistic stimulus-dependent correlational contribution quantifying the information content of changes of correlations with stimulus, and a stimulus-independent correlational contribution term that reflects interactions between the distribution of rates of individual cells and the average level of cross-correlation. We apply the new method to simultaneously recorded data from somatosensory and visual cortices. We demonstrate that it constitutes a reliable tool to determine the role of cross-correlated activity in stimulus coding even when high firing rate data (such as multi-unit recordings) are considered.     

Compound-specific carbon isotope analysis of volatile organic compounds in the low-microgram per liter range

Compound-specific carbon isotope analysis (CSIA) has become an important tool in biological, archeological, and geological studies as well as in forensics, food sciences, and organic chemistry. If sensitivity could be enhanced, CSIA would further have an improved potential for environmental applications such as, for example, in situ remediation studies to assess contaminated environments, identification of pollutant degradation pathways and kinetics, distinction between degradation/formation mechanisms, or, verification of contaminant sources. With this goal in mind, we have developed methods to determine delta13C values of commonly reported groundwater contaminants in low-microgram per liter concentrations. Several injection and preconcentration techniques were evaluated for this purpose, i.e., on-column injection, split/ splitless injection, solid-phase microextraction (SPME), and purge and trap (P&T) in combination with gas chromatography-isotope ratio mass spectrometry. The delta13C values of the target compounds were determined by liquid injections of the analytes dissolved in diethyl ether or, in the case of P&T and SPME, by extraction from water spiked with the analytes. P&T extraction was the most efficient preconcentration technique reaching method detection limits (MDLs) from 0.25 to 5.0 microg/L. These are the lowest MDLs reported so far for continuous-flow isotope ratio determinations, using a commercially available and fully automated system. Isotopic fractionation resulting from preconcentration and injection was investigated and quantified for the priority groundwater pollutants methyl tert-butyl ether (MTBE), chloroform, tetrachloromethane, chlorinated ethylenes, benzene, and toluene. The isotopic fractionations caused by the extraction techniques were small but highly reproducible and could therefore be corrected for. P&T was characterized by a higher reproducibility and smaller isotopic fractionations than SPME. Among the liquid injection techniques, cold on-column injection resulted in slightly better precision compared to split/splitless injection. However, the MDLs determined for liquid injections were 4-6 orders of magnitude higher (i.e., 9.5-2800 mg/L) than for P&T and SPME. Since both of the latter methods are solventless, a better chromatographic resolution was obtained than for the liquid injection techniques. The P&T and SPME methods described here are also applicable for CSIA of D/H ratios, which require 10-20 times higher analyte concentrations than 13C/12C analysis. Finally, the applicability of the described methods is demonstrated for pollutant concentrations of only 5-60 microg/L in environmental samples.     

Analytical methods for thorium determination: a journey from conventional methods to novel applications

Thorium may be determined using a variety of analytical methods. These may be based on chemical or physical principles or may make use of the radioactive decay of the individual thorium isotopes. The method to be used for a certain analytical purpose is selected as a function of the matrix of the sample and of the concentration of thorium. The present paper describes different methodologies, their range of application and discusses selected results. The methods described cover a concentration range of more than twelve orders of magnitude, i.e. from concentrated solutions to micrometre-sized particles. Emphasis is given to active radiometric techniques, chemical and instrumental analysis.     

Errors-in-variables methods in system identification

The paper gives a survey of errors-in-variables methods in system identification. Background and motivation are given, and examples illustrate why the identification problem can be difficult. Under general weak assumptions, the systems are not identifiable, but can be parameterized using one degree-of-freedom. Examples where identifiability is achieved under additional assumptions are also provided. A number of approaches for parameter estimation of errors-in-variables models are presented. The underlying assumptions and principles for each approach are highlighted.