A novel monoclonal antibody suitable for the detection of leukotriene B4.

Leukotriene B4 as an inflammatory mediator is an important biomarker for different respiratory diseases like asthma, chronic obstructive pulmonary disease or cystic lung fibrosis. Therefore the detection of LTB4 is helpful in the diagnosis of these pulmonary diseases. However, until now its determination in exhaled breath condensates suffers from problems of accuracy.

Label free sensing of creatinine using a 6 GHz CMOS near-field dielectric immunosensor.

In this work we present a CMOS high frequency direct immunosensor operating at 6 GHz (C-band) for label free determination of creatinine. The sensor is fabricated in standard 0.13 μm SiGe:C BiCMOS process.

Point-of-care testing of proteins.

Point-of-care testing (POCT) is a fast developing area in clinical diagnostics that is considered to be one of the main driving forces for the future in vitro diagnostic market. POCT means decentralized testing at the site of patient care. The most important POCT devices are handheld blood glucose sensors.

Analysis of recognition of fructose by imprinted polymers.

Binding of fructose to the fructose imprinted polymer (MIP(Frc)) and pinacol imprinted polymer (control) were studied both in batch and a flow through mode. The influence of the cross-linkers ethylene glycol dimethacrylate (EDMA) and trimethylolpropane trimethacrylate (TRIM) on the binding characteristics was analysed. TRIM cross-linked MIPs showed a lower (unspecific) binding for the control polymer (pinacol imprinted) and higher binding of fructose as compared with the EDMA-MIPs.

Sequential conversion by catalytically active MIP and immobilized tyrosinase in a thermistor.

To amplify the heat-signal generated by MIP catalyzed solvolysis of phenylacetate the reaction has been combined for the first time in a reactor with the subsequent oxidation by immobilized tyrosinase. The polymer cleaves the substrate and the released phenol is afterwards converted to o-benzoquinone by the tyrosinase. The separated and sequentially coupled reactions are characterized by the heat generated in a thermistor.

Thermometric MIP sensor for fructosyl valine.

Interactions of molecularly imprinted polymers containing phenyl boronic acid residues with fructosyl valine, fructose and pinacol, respectively are analysed in aqueous solution (pH 11.4) by using a flow calorimeter. The reversible formation of (two) cyclic boronic acid diesters per fructosyl molecule generates a 40-fold higher exothermic signal as compared to the control polymer.

Electrochemical biochips for protein analysis.

Proteins bear important functions for most life processes. It is estimated that the human proteome comprises more than 250,000 proteins. Over the last years, highly sophisticated and powerful instruments have been developed that allow their detection and characterization with great precision and sensitivity.

Development of a small peptide tag for covalent labeling of proteins.

A 21-mer peptide that can be used to covalently introduce synthetic molecules into proteins has been developed. Phage-displayed peptide libraries were subjected to reaction-based selection with 1,3-diketones. The peptide was further evolved by addition of a randomized region and reselection for improved binding.

Enzyme activity control by responsive redoxpolymers.

A new thermoresponsive poly-N-isopropylacrylamide (PNIPAM)-ferrocene polymer was synthesized and characterized. PNIPAMFoxy bears additional oxirane groups which were used for attachment by a self-assembly process on a cysteamine-modified gold electrode to create a thin hydrophilic film. The new redox polymer enabled electrical communication between the cofactor pyrrolinoquinoline quinone (PQQ) of soluble glucose dehydrogenase (sGDH) and the electrode for sensitive detection of this enzyme as a prospective protein label.

Development of fructosyl valine binding polymers by covalent imprinting.

Molecularly imprinted polymers (MIPs) against fructosyl valine (Fru-Val), the N-terminal constituent of hemoglobin A1c beta-chains, were prepared by cross-linking of beta-D-Fru-Val-O-bis(4-vinylphenylboronate) with an excess of ethylene glycol dimethacrylate (EDMA) or trimethylolpropane trimethacrylate (TRIM). Control MIPs were prepared in analogy by cross-linking the corresponding vinylphenylboronate esters of fructose and pinacol. After template extraction batch rebinding studies were performed using different pH values and buffer compositions.

Homogeneous indirect fluorescence quenching immunoassay for the determination of low molecular weight substances.

This paper describes the principle of a homogeneous indirect fluorescence quenching immunoassay that uses monoclonal antibodies. It is a carrier-free assay system that is performed completely in solution. The assay system was established for the determination of a low molecular weight substance (hapten), the herbicide diuron, used as a model analyte.

Affinity interactions between phenylboronic acid-carrying self-assembled monolayers and flavin adenine dinucleotide or horseradish peroxidase.

A method is provided for the recognition of glycated molecules based on their binding affinities to boronate-carrying monolayers. The affinity interaction of flavin adenine dinucleotide (FAD) and horseradish peroxidase (HRP) with phenylboronic acid monolayers on gold was investigated by using voltammetric and microgravimetric methods. Conjugates of 3-aminophenylboronic acid and 3,3'-dithiodipropionic acid di(N-hydroxysuccinimide ester) or 11-mercaptoundecanoic acid were prepared and self-assembled on gold surfaces to generate monolayers.

Electrochemical bioassay utilizing encapsulated electrochemical active microcrystal biolabels.

A new approach to perform electrochemical immunoassay based on the utilization of encapsulated microcrystal was developed. The microcrystal labels create a "supernova effect" upon exposure to a desired releasing agent. The microcrystal cores dissolve, and large amounts of signal-generating molecules diffuse across the capsule wall into the outer environment.

Detection of subpicomolar concentrations of human matrix metalloproteinase-2 by an optical biosensor.

We describe in this paper the development of a one-step sandwich assay for the highly sensitive and fast detection of human matrix metalloproteinase (MMP)-2 (EC 3.4.24.

Analysis of thiols with tyrosinase-modified carbon paste electrodes based on blocking of substrate recycling.

The enzyme, tyrosinase, was immobilized inside carbon paste electrodes (CPE) for the analysis of thiol-containing compounds such as the reduced form of glutathione (GSH) and L-cysteine. The measuring principle of this sensor is based on the blocking of the substrate recycling process between the enzyme and the electrode. The current response is monitored at -0.

Development of a flow-injection analysis (FIA) enzyme sensor for fructosyl amine monitoring.

An enzyme-sensor system with flow-injection analysis (FIA) has been developed for the detection of fructosyl amine compounds; the sensor utilizes fructosyl amine oxidase isolated from the marine yeast Pichia sp. N1-1 strain. With this FIA system 0.

Activation of cellulose membranes with 1,1'-carbonyldiimidazole or 1-cyano-4-dimethylaminopyridinium tetrafluoroborate as a basis for the development of immunosensors.

Methods for the activation of a cellulose dialysis membrane for immunosensor applications have been developed. For activation two reagents, 1,1'-carbonyldiimidazole (CDI) and 1-cyano-4-dimethylaminopyridinium tetrafluoroborate (CDAP), were compared with respect to the coupling efficiency for glucose oxidase (GOx) and 1,8-diamino-2,6-dioxaoctane. The maximum level of activation was 2.

Electrochemical immunoassays.

Immunoassays (IA) use the specific antigen antibody complexation for analytical purposes. Radioimmunoassays (RIA), fluorescence immunoassays (FIA) and enzyme immunoassays (EIA) are well established in clinical diagnostics. For the development of hand-held devices which can be used for point of care measurements, electrochemical immunoassays are promising alternatives to existing immunochemical tests.

Enzyme kinetic assays with surface plasmon resonance (BIAcore) based on competition between enzyme and creatinine antibody.

A procedure is described which allows the characterization of enzyme by a hybrid approach using an enzyme and an antibody. The presented method is related to the affinity determination of antibodies by the 'affinity in solution' procedure for BlAcore. The antibody is used as an indicator for the concentration of substrate, which is also the antigen.

Research and development in biosensors.

Progress in biosensors has mainly been made by the improvement of the biological components and the implementation of microsystem technologies. Enzymes are still the most appropriate recognition elements because they combine high chemical specificity and inherent biocatalytic signal amplification. A breakthrough has been achieved in the application of membrane-integrated receptor systems for analyte recognition and signal transduction in biosensors.

Operation of a miniature redox hydrogel-based pyruvate sensor in undiluted deoxygenated calf serum.

An amperometric sensor for the detection of pyruvate in biological fluids was formed by modifying the tip of a 0.25 mm gold wire with a layer of electrically "wired" recombinant pyruvate oxidase (POP). The sensor did not require O2 for its operation.

Development of a creatinine ELISA and an amperometric antibody-based creatine sensor with a detection limit in the nanomolar range.

Creatinine-specific antibodies have been generated and used for highly sensitive and specific immunochemical creatinine determinations. Creatinine was derivatized at N3 and coupled to KLH carrier protein. On the basis of this immunogen, monoclonal antibodies were developed by hybridoma technology.

Production and characterization of monoclonal antibodies against urea derivatives.

A panel of monoclonal antibodies was generated against the urea-based hapten N-(2-N-chloroacetylaminobenzyl)-N'-4-chlorophenylurea as a tool for building up sensitive immune assays to detect urea derivatives and to screen them for catalytic antibodies (Abs). Eleven hybridomas were obtained that produced Abs reactive to the hapten. All Abs were of IgG class.