Refine
Year of publication
Document Type
- Article (540)
- Conference Proceeding (52)
- Part of a Book (9)
- Doctoral Thesis (4)
- Book (3)
- Other (2)
- Report (2)
- Patent (1)
Language
- English (555)
- German (57)
- Multiple languages (1)
Keywords
- Biosensor (6)
- Graduiertentagung (4)
- biosensors (4)
- frequency mixing magnetic detection (4)
- LAPS (3)
- Label-free detection (3)
- Light-addressable potentiometric sensor (3)
- capacitive field-effect sensor (3)
- field-effect sensor (3)
- hydrogen peroxide (3)
- magnetic nanoparticles (3)
- tobacco mosaic virus (TMV) (3)
- Acyl-amino acids (2)
- Aminoacylase (2)
- Bacillaceae (2)
- Bacillus atrophaeus (2)
- Biotechnological application (2)
- Calorimetric gas sensor (2)
- Capacitive field-effect sensor (2)
- Hydrogen peroxide (2)
Institute
- INB - Institut für Nano- und Biotechnologien (613) (remove)
Biomedical applications of magnetic nanoparticles (MNP) fundamentally rely on the particles’ magnetic relaxation as a response to an alternating magnetic field. The magnetic relaxation complexly depends on the interplay of MNP magnetic and physical properties with the applied field parameters. It is commonly accepted that particle core size is a major contributor to signal generation in all the above applications, however, most MNP samples comprise broad distribution spanning nm and more. Therefore, precise knowledge of the exact contribution of individual core sizes to signal generation is desired for optimal MNP design generally for each application. Specifically, we present a magnetic relaxation simulation-driven analysis of experimental frequency mixing magnetic detection (FMMD) for biosensing to quantify the contributions of individual core size fractions towards signal generation. Applying our method to two different experimental MNP systems, we found the most dominant contributions from approx. 20 nm sized particles in the two independent MNP systems. Additional comparison between freely suspended and immobilized MNP also reveals insight in the MNP microstructure, allowing to use FMMD for MNP characterization, as well as to further fine-tune its applicability in biosensing.
Persistent infection with high-risk human papillomaviruses (hrHPV) can result in the formation of anogenital cancers. As hrHPV proteins E6 and E7 are required for cancer initiation and maintenance, they are ideal targets for immunotherapeutic interventions. Previously, we have described the development of DNA vaccines for the induction of HPV16 E6 and E7 specific T cell immunity. These vaccines consist of ‘gene-shuffled’ (SH) versions of HPV16 E6 and E7 that were fused to Tetanus Toxin Fragment C domain 1 (TTFC) and were named TTFC-E6SH and TTFC-E7SH. Gene-shuffling was performed to avoid the risk of inducing malignant transformation at the vaccination site. Here, we describe the preclinical safety evaluation of these candidate vaccines by analysis of their transforming capacity in vitro using established murine fibroblasts (NIH 3T3 cells) and primary human foreskin keratinocytes (HFKs). We demonstrate that neither ectopic expression of TTFC-E6SH and TTFC-E7SH alone or in combination enabled NIH 3T3 cells to form colonies in soft agar. In contrast, expression of HPV16 E6WT and E7WT alone or in combination resulted in effective transformation. Similarly, retroviral transduction of HFKs from three independent donors with both TTFC-E6SH and TTFC-E7SH alone or in combination did not show any signs of immortalization. In contrast, the combined expression of E6WT and E7WT induced immortalization in HFKs from all donors. Based on these results we consider it justified to proceed to clinical evaluation of DNA vaccines encoding TTFC-E6SH and TTFC-E7SH in patients with HPV16 associated (pre)malignancies.
Planar and three-dimensional (3D) interdigitated electrodes (IDE) with electrode digits separated by an insulating barrier of different heights were electrochemically characterized and compared in terms of their sensing properties. Due to the impact of the surface resistance, both types of IDE structures display a non-linear behavior in low-ionic strength solutions. The experimental data were fitted to an electrical equivalent circuit and interpreted taking into account the surface-charge-governed properties. The effect of a charged polyelectrolyte layer electrostatically assembled onto the sensor surface on the surface resistance in solutions with different KCl concentration is studied. In case of the same electrode footprint, 3D-IDEs show a larger cell constant and a higher sensitivity to molecular adsorption than that of planar IDEs. The obtained results demonstrate the potential of 3D-IDEs as a new transducer structure for a direct label-free sensing of charged molecules.
The subtilase family (S8), a member of the clan SB of serine proteases are ubiquitous in all kingdoms of life and fulfil different physiological functions. Subtilases are divided in several groups and especially subtilisins are of interest as they are used in various industrial sectors. Therefore, we searched for new subtilisin sequences of the family Bacillaceae using a data mining approach. The obtained 1,400 sequences were phylogenetically classified in the context of the subtilase family. This required an updated comprehensive overview of the different groups within this family. To fill this gap, we conducted a phylogenetic survey of the S8 family with characterised holotypes derived from the MEROPS database. The analysis revealed the presence of eight previously uncharacterised groups and 13 subgroups within the S8 family. The sequences that emerged from the data mining with the set filter parameters were mainly assigned to the subtilisin subgroups of true subtilisins, high-alkaline subtilisins, and phylogenetically intermediate subtilisins and represent an excellent source for new subtilisin candidates.
In this study we show an optical biosensor concept, based on elastic light scattering from sapphire microspheres. Transmitted and elastic scattering intensity of the microspheres (radius 500 μm, refractive index 1.77) on an optical fiber half coupler is analyzed at 1510 nm. The 0.43 nm angular mode spacing of the resonances is comparable to the angular mode spacing value estimated using the optical size of the microsphere. The spectral linewidths of the resonances are in the order of 0.01 nm, which corresponds to quality factors of approximately 105. A polydopamine layer is used as a functionalizing agent on sapphire microspherical resonators in view of biosensor implementation. The varying layer thickness on the microsphere is determined as a function of the resonance wavelength shift. It is shown that polymer functionalization has a minor effect on the quality factor. This is a promising step toward the development of an optical biosensor.
Photoelectrochemical (PEC) biosensors are a rather novel type of biosensors thatutilizelighttoprovideinformationaboutthecompositionofananalyte,enablinglight-controlled multi-analyte measurements. For enzymatic PEC biosensors,amperometric detection principles are already known in the literature. In con-trast, there is only a little information on H+-ion sensitive PEC biosensors. Inthis work, we demonstrate the detection of H+ions emerged by H+-generatingenzymes, exemplarily demonstrated with penicillinase as a model enzyme on atitanium dioxide photoanode. First, we describe the pH sensitivity of the sensorand study possible photoelectrocatalytic reactions with penicillin. Second, weshow the enzymatic PEC detection of penicillin.
The presentation of enzymes on viral scaffolds has beneficial effects such as an increased enzyme loading and a prolonged reusability in comparison to conventional immobilization platforms. Here, we used modified tobacco mosaic virus (TMV) nanorods as enzyme carriers in penicillin G detection for the first time. Penicillinase enzymes were conjugated with streptavidin and coupled to TMV rods by use of a bifunctional biotin-linker. Penicillinase-decorated TMV particles were characterized extensively in halochromic dye-based biosensing. Acidometric analyte detection was performed with bromcresol purple as pH indicator and spectrophotometry. The TMV-assisted sensors exhibited increased enzyme loading and strongly improved reusability, and higher analysis rates compared to layouts without viral adapters. They extended the half-life of the sensors from 4 - 6 days to 5 weeks and thus allowed an at least 8-fold longer use of the sensors. Using a commercial budget-priced penicillinase preparation, a detection limit of 100 µM penicillin was obtained. Initial experiments also indicate that the system may be transferred to label-free detection layouts.
Penicillin detection by means of field-effect based sensors: EnFET, capacitive EIS sensor or LAPS?
(2001)
Penicillin detection by means of field-effect based sensors: EnFET, capacitive EIS sensor or LAPS?
(2000)
Optoelectronic Properties of Nanostructured Ensembles Controlled by Biomolecular Logic Systems
(2008)
Optimization of the immobilization of bacterial spores on glass substrates with organosilanes
(2016)
Spores can be immobilized on biosensors to function as sensitive recognition elements. However, the immobilization can affect the sensitivity and reproducibility of the sensor signal. In this work, three different immobilization strategies with organosilanes were optimized and characterized to immobilize Bacillus atrophaeus spores on glass substrates. Five different silanization parameters were investigated: nature of the solvent, concentration of the silane, silanization time, curing process, and silanization temperature. The resulting silane layers were resistant to a buffer solution (e.g., Ringer solution) with a polysorbate (e.g., Tween®80) and sonication.
Bacillus subtilis and Bacillus licheniformis are widely used for the large-scale industrial production of proteins. These strains can efficiently secrete proteins into the culture medium using the general secretion (Sec) pathway. A characteristic feature of all secreted proteins is their N-terminal signal peptides, which are recognized by the secretion machinery. Here, we have studied the production of an industrially important secreted protease, namely, subtilisin BPN′ from Bacillus amyloliquefaciens. One hundred seventy-three signal peptides originating from B. subtilis and 220 signal peptides from the B. licheniformis type strain were fused to this secretion target and expressed in B. subtilis, and the resulting library was analyzed by high-throughput screening for extracellular proteolytic activity. We have identified a number of signal peptides originating from both organisms which produced significantly increased yield of the secreted protease. Interestingly, we observed that levels of extracellular protease were improved not only in B. subtilis, which was used as the screening host, but also in two different B. licheniformis strains. To date, it is impossible to predict which signal peptide will result in better secretion and thus an improved yield of a given extracellular target protein. Our data show that screening a library consisting of homologous and heterologous signal peptides fused to a target protein can identify more-effective signal peptides, resulting in improved protein export not only in the original screening host but also in different production strains.
Optimization of passivation layers for corrosion protection of silicon-based microelectrode arrays
(2000)
A light-addressable potentiometric sensor (LAPS) is a field-effect-based potentiometric device, which detects concentration changes of an analyte solution on the sensor surface in a spatially resolved way. It uses a light source to generate electron–hole pairs inside the semiconductor, which are separated in the depletion region due to an applied bias voltage across the sensor structure and hence, a surface-potential-dependent photocurrent can be read out. However, depending on the beam angle of the light source, scattering effects can occur, which influence the recorded signal in LAPS-based differential measurements. To solve this problem, a novel illumination unit based on a field programmable gate array (FPGA) consisting of 16 small-sized tunable infrared laser-diode modules (LDMs) is developed. Due to the improved focus of the LDMs with a beam angle of only 2 mrad, undesirable scattering effects are minimized. Escherichia coli (E. coli) K12 bacteria are used as a test microorganism to study the extracellular acidification on the sensor surface. Furthermore, a salt bridge chamber is built up and integrated with the LAPS system enabling multi-chamber differential measurements with a single Ag/AgCl reference electrode.
The immobilization of NAD+-dependent dehydrogenases, in combination with a diaphorase, enables the facile development of multiparametric sensing devices. In this work, an amperometric biosensor array for simultaneous determination of ethanol, formate, d- and l-lactate is presented. Enzyme immobilization on platinum thin-film electrodes was realized by chemical cross-linking with glutaraldehyde. The optimization of the sensor performance was investigated with regard to enzyme loading, glutaraldehyde concentration, pH, cofactor concentration and temperature. Under optimal working conditions (potassium phosphate buffer with pH 7.5, 2.5 mmol L-1 NAD+, 2.0 mmol L-1 ferricyanide, 25 °C and 0.4% glutaraldehyde) the linear working range and sensitivity of the four sensor elements was improved. Simultaneous and cross-talk free measurements of four different metabolic parameters were performed successfully. The reliable analytical performance of the biosensor array was demonstrated by application in a clarified sample of inoculum sludge. Thereby, a promising approach for on-site monitoring of fermentation processes is provided.
Online-Messsysteme für die automatisierte Charakterisierung von feldeffektbasierten Biosensoren
(2007)
A variety of transition metals, e.g., copper, zinc, cadmium, lead, etc. are widely used in industry as components for wires, coatings, alloys, batteries, paints and so on. The inevitable presence of transition metals in industrial processes implies the ambition of developing a proper analytical technique for their adequate monitoring. Most of these elements, especially lead and cadmium, are acutely toxic for biological organisms. Quantitative determination of these metals at low activity levels in different environmental and industrial samples is therefore a vital task. A promising approach to achieve an at-side or on-line monitoring on a miniaturized and cost efficient way is the combination of a common potentiometric sensor array with heavy metal-sensitive thin-film materials, like chalcogenide glasses and polymeric materials, respectively.
N-Acyl-amino acids can act as mild biobased surfactants, which are used, e.g., in baby shampoos. However, their chemical synthesis needs acyl chlorides and does not meet sustainability criteria. Thus, the identification of biocatalysts to develop greener synthesis routes is desirable. We describe a novel aminoacylase from Paraburkholderia monticola DSM 100849 (PmAcy) which was identified, cloned, and evaluated for its N-acyl-amino acid synthesis potential. Soluble protein was obtained by expression in lactose autoinduction medium and co-expression of molecular chaperones GroEL/S. Strep-tag affinity purification enriched the enzyme 16-fold and yielded 15 mg pure enzyme from 100 mL of culture. Biochemical characterization revealed that PmAcy possesses beneficial traits for industrial application like high temperature and pH-stability. A heat activation of PmAcy was observed upon incubation at temperatures up to 80 °C. Hydrolytic activity of PmAcy was detected with several N-acyl-amino acids as substrates and exhibited the highest conversion rate of 773 U/mg with N-lauroyl-L-alanine at 75 °C. The enzyme preferred long-chain acyl-amino-acids and displayed hardly any activity with acetyl-amino acids. PmAcy was also capable of N-acyl-amino acid synthesis with good conversion rates. The best synthesis results were obtained with the cationic L-amino acids L-arginine and L-lysine as well as with L-leucine and L-phenylalanine. Exemplarily, L-phenylalanine was acylated with fatty acids of chain lengths from C8 to C18 with conversion rates of up to 75%. N-lauroyl-L-phenylalanine was purified by precipitation, and the structure of the reaction product was verified by LC–MS and NMR.
A novel photoexcitation method for the light-addressable potentiometric sensor (LAPS) is proposed to achieve a higher spatial resolution of chemical images. The proposed method employs a combined light source that consists of a modulated light probe, which generates the alternating photocurrent signal, and a ring of constant illumination surrounding it. The constant illumination generates a sheath of carriers with increased concentration which suppresses the spread of photocarriers by enhanced recombination. A device simulation was carried out to verify the effect of constant illumination on the spatial resolution, which demonstrated that a higher spatial resolution can be obtained.
Novel organic membrane-based thin-film microsensors for the determination of heavy metal cations
(2006)
A first step towards the fabrication and electrochemical evaluation of thin-film microsensors based on organic PVC membranes for the determination of Hg(II), Cd(II), Pb(II) and Cu(II) ions in solutions has been realised. The membrane-coating mixture used in the preparation of this new type of microsensors is incorporating PVC as supporting matrix, o-nitrophenyloctylether (o-NPOE) as solvent mediator and a recently synthesized Hg[dimethylglyoxime(phene)]2+ and Bis-(4-hydroxyacetophenone)-ethylenediamine as electroactive materials for Hg(II) and Cd(II), respectively. A set of three commercialised ionophores for Cd(II), Pb(II) and Cu(II) has been also used for comparison. Thin-film microsensors based on these membranes showed a Nernstian response of slope (26-30 mV/dec.) for the respective tested cations. The potentiometric response characteristics (linear range, pH range, detection limit and response time) are comparable with those obtained by conventional membranes as well as coated wire electrodes prepared from the same membrane. The realisation of the new organic membrane-based thin-film microsensors overcomes the problem of an insufficient selectivity of solid-state-based thinfilm sensors.