TY - JOUR A1 - Laux, Eva-Maria A1 - Bier, Frank Fabian A1 - Hölzel, Ralph T1 - Electrode-based AC electrokinetics of proteins BT - a mini-review JF - Bioelectrochemistry : official journal of the Bioelectrochemical Society ; an international journal devoted to electrochemical aspects of biology and biological aspects of electrochemistry N2 - Employing electric phenomena for the spatial manipulation of bioparticles from whole cells down to dissolved molecules has become a useful tool in biotechnology and analytics. AC electrokinetic effects like dielectrophoresis and AC electroosmosis are increasingly used to concentrate, separate and immobilize DNA and proteins. With the advance of photolithographical micro- and nanofabrication methods, novel or improved bioanalytical applications benefit from concentrating analytes, signal enhancement and locally controlled immobilization by AC electrokinetic effects. In this review of AC electrokinetics of proteins, the respective studies are classified according to their different electrode geometries: individual electrode pairs, interdigitated electrodes, quadrupole electrodes, and 3D configurations of electrode arrays. Known advantages and disadvantages of each layout are discussed. KW - AC electrokinetics KW - Dielectrophoresis KW - Electrodes KW - Electroosmosis KW - Proteins Y1 - 2017 U6 - https://doi.org/10.1016/j.bioelechem.2017.11.010 SN - 1567-5394 SN - 1878-562X VL - 120 SP - 76 EP - 82 PB - Elsevier B.V. CY - Amsterdam ER - TY - JOUR A1 - Kersting, Sebastian A1 - Rausch, Valentina A1 - Bier, Frank Fabian A1 - von Nickisch-Rosenegk, Markus T1 - A recombinase polymerase amplification assay for the diagnosis of atypical pneumonia JF - Analytical biochemistry : methods in the biological sciences N2 - Pneumonia is one of the most common and potentially lethal infectious conditions worldwide. Streptococcus pneumoniae is the pathogen most frequently associated with bacterial community-acquired pneumonia, while Legionella pneumophila is the major cause for local outbreaks of legionellosis. Both pathogens can be difficult to diagnose since signs and symptoms are nonspecific and do not differ from other causes of pneumonia. Therefore, a rapid diagnosis within a clinically relevant time is essential for a fast onset of the proper treatment. Although methods based on polymerase chain reaction significantly improved the identification of pathogens, they are difficult to conduct and need specialized equipment. We describe a rapid and sensitive test using isothermal recombinase polymerase amplification and detection on a disposable test strip. This method does not require any special instrumentation and can be performed in less than 20 min. The analytical sensitivity in the multiplex assay amplifying specific regions of S. pneumoniae and L. pneumophila simultaneously was 10 CFUs of genomic DNA per reaction. In cross detection studies with closely related strains and other bacterial agents the specificity of the RPA was confirmed. The presented method is applicable for near patient and field testing with a rather simple routine and the possibility for a read out with the naked eye. Y1 - 2018 U6 - https://doi.org/10.1016/j.ab.2018.04.014 SN - 0003-2697 SN - 1096-0309 VL - 550 SP - 54 EP - 60 PB - Elsevier CY - San Diego ER - TY - GEN A1 - Breitenstein, Michael A1 - Nielsen, Peter E. A1 - Hölzel, Ralph A1 - Bier, Frank Fabian T1 - DNA-nanostructure-assembly by sequential spotting T2 - Postprints der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe N2 - Background: The ability to create nanostructures with biomolecules is one of the key elements in nanobiotechnology. One of the problems is the expensive and mostly custom made equipment which is needed for their development. We intended to reduce material costs and aimed at miniaturization of the necessary tools that are essential for nanofabrication. Thus we combined the capabilities of molecular ink lithography with DNA-self-assembling capabilities to arrange DNA in an independent array which allows addressing molecules in nanoscale dimensions. Results: For the construction of DNA based nanostructures a method is presented that allows an arrangement of DNA strands in such a way that they can form a grid that only depends on the spotted pattern of the anchor molecules. An atomic force microscope (AFM) has been used for molecular ink lithography to generate small spots. The sequential spotting process allows the immobilization of several different functional biomolecules with a single AFM-tip. This grid which delivers specific addresses for the prepared DNA-strand serves as a two-dimensional anchor to arrange the sequence according to the pattern. Once the DNA-nanoarray has been formed, it can be functionalized by PNA (peptide nucleic acid) to incorporate advanced structures. Conclusions: The production of DNA-nanoarrays is a promising task for nanobiotechnology. The described method allows convenient and low cost preparation of nanoarrays. PNA can be used for complex functionalization purposes as well as a structural element. T3 - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 1027 KW - atomic force microscope KW - peptide nucleic acid KW - persistence length KW - adapter oligonucleotide KW - high fluorescence signal Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-431108 SN - 1866-8372 IS - 1027 ER - TY - JOUR A1 - Kagel, Heike A1 - Bier, Frank Fabian A1 - Frohme, Marcus A1 - Glökler, Jörn F. T1 - A Novel Optical Method To Reversibly Control Enzymatic Activity Based On Photoacids JF - Scientific reports N2 - Most biochemical reactions depend on the pH value of the aqueous environment and some are strongly favoured to occur in an acidic environment. A non-invasive control of pH to tightly regulate such reactions with defined start and end points is a highly desirable feature in certain applications, but has proven difficult to achieve so far. We report a novel optical approach to reversibly control a typical biochemical reaction by changing the pH and using acid phosphatase as a model enzyme. The reversible photoacid G-acid functions as a proton donor, changing the pH rapidly and reversibly by using high power UV LEDs as an illumination source in our experimental setup. The reaction can be tightly controlled by simply switching the light on and off and should be applicable to a wide range of other enzymatic reactions, thus enabling miniaturization and parallelization through non-invasive optical means. Y1 - 2019 U6 - https://doi.org/10.1038/s41598-019-50867-w SN - 2045-2322 VL - 9 PB - Nature Publishing Group CY - London ER - TY - GEN A1 - Stanke, S. A1 - Wenger, C. A1 - Bier, Frank Fabian A1 - Hölzel, Ralph T1 - Dielectrophoretic functionalization of nanoelectrode arrays for the detection of influenza viruses T2 - European biophysics journal : with biophysics letters ; an international journal of biophysics Y1 - 2017 SN - 0175-7571 SN - 1432-1017 VL - 46 SP - S337 EP - S337 PB - Springer CY - New York ER - TY - GEN A1 - Laux, Eva-Maria A1 - Gibbons, J. A1 - Ermilova, Elena A1 - Bier, Frank Fabian A1 - Hölzel, Ralph T1 - Broadband dielectric spectroscopy of bovine serum albumin in the GHz range T2 - European biophysics journal : with biophysics letters ; an international journal of biophysics Y1 - 2017 SN - 0175-7571 SN - 1432-1017 VL - 46 SP - S347 EP - S347 PB - Springer CY - New York ER - TY - GEN A1 - Laux, Eva-Maria A1 - Knigge, Xenia A1 - Wenger, C. A1 - Bier, Frank Fabian A1 - Hölzel, Ralph T1 - AC electrokinetic manipulation of nanoparticles and molecules T2 - European biophysics journal : with biophysics letters ; an international journal of biophysics Y1 - 2017 SN - 0175-7571 SN - 1432-1017 VL - 46 SP - S189 EP - S189 PB - Springer CY - New York ER - TY - GEN A1 - Knigge, Xenia A1 - Wenger, C. A1 - Bier, Frank Fabian A1 - Hölzel, Ralph T1 - AC electrokinetic immobilisation of nanoobjects as individual singles in regular arrays T2 - European biophysics journal : with biophysics letters ; an international journal of biophysics Y1 - 2017 SN - 0175-7571 SN - 1432-1017 VL - 46 SP - S187 EP - S187 PB - Springer CY - New York ER - TY - JOUR A1 - Connor, Daniel Oliver A1 - Danckert, Lena A1 - Hoppe, Sebastian A1 - Bier, Frank Fabian A1 - von Nickisch-Rosenegk, Markus T1 - Epitope determination of immunogenic proteins of Neisseria gonorrhoeae JF - PLoS one N2 - Neisseria gonorrhoeae is the causative organism of gonorrhoea, a sexually transmitted disease that globally accounts for an estimated 80 to 100 million new infections per year. Increasing resistances to all common antibiotics used for N. gonorrhoeae treatment pose the risk of an untreatable disease. Further knowledge of ways of infection and host immune response are needed to understand the pathogen-host interaction and to discover new treatment alternatives against this disease. Therefore, detailed information about immunogenic proteins and their properties like epitope sites could advance further research in this area. In this work, we investigated immunogenic proteins of N. gonorrhoeae for linear epitopes by microarrays. Dominant linear epitopes were identified for eleven of the nineteen investigated proteins with three polyclonal rabbit antibodies from different immunisations. Identified linear epitopes were further examined for non-specific binding with antibodies to Escherichia coli and the closely related pathogen Neisseria meningitidis. On top of that, amino acids crucial for the antibody epitope binding were detected by microarray based alanine scans. Y1 - 2017 U6 - https://doi.org/10.1371/journal.pone.0180962 SN - 1932-6203 VL - 12 PB - PLoS CY - San Fransisco ER - TY - JOUR A1 - Bader, Denise A1 - Klier, Dennis Tobias A1 - Hettrich, C. A1 - Bier, Frank Fabian A1 - Wessig, Pablo T1 - Detecting carbohydrate-lectin interactions using a fluorescent probe based on DBD dyes JF - Analytical methods : advancing methods and applications N2 - Herein we present an efficient synthesis of a biomimetic probe with modular construction that can be specifically bound by the mannose binding FimH protein - a surface adhesion protein of E. coli bacteria. The synthesis combines the new and interesting DBD dye with the carbohydrate ligand mannose via a Click reaction. We demonstrate the binding to E. coli bacteria over a large concentration range and also present some special characteristics of those molecules that are of particular interest for the application as a biosensor. In particular, the mix-and-measure ability and the very good photo-stability should be highlighted here. Y1 - 2016 U6 - https://doi.org/10.1039/c5ay02991k SN - 1759-9660 SN - 1759-9679 VL - 8 SP - 1235 EP - 1238 PB - Royal Society of Chemistry CY - Cambridge ER - TY - JOUR A1 - Connor, Daniel Oliver A1 - Zantow, Jonas A1 - Hust, Michael A1 - Bier, Frank Fabian A1 - von Nickisch-Rosenegk, Markus T1 - Identification of Novel Immunogenic Proteins of Neisseria gonorrhoeae by Phage Display JF - PLoS one N2 - Neisseria gonorrhoeae is one of the most prevalent sexually transmitted diseases worldwide with more than 100 million new infections per year. A lack of intense research over the last decades and increasing resistances to the recommended antibiotics call for a better understanding of gonococcal infection, fast diagnostics and therapeutic measures against N. gonorrhoeae. Therefore, the aim of this work was to identify novel immunogenic proteins as a first step to advance those unresolved problems. For the identification of immunogenic proteins, pHORF oligopeptide phage display libraries of the entire N. gonorrhoeae genome were constructed. Several immunogenic oligopeptides were identified using polyclonal rabbit antibodies against N. gonorrhoeae. Corresponding full-length proteins of the identified oligopeptides were expressed and their immunogenic character was verified by ELISA. The immunogenic character of six proteins was identified for the first time. Additional 13 proteins were verified as immunogenic proteins in N. gonorrhoeae. Y1 - 2016 U6 - https://doi.org/10.1371/journal.pone.0148986 SN - 1932-6203 VL - 11 PB - PLoS CY - San Fransisco ER - TY - JOUR A1 - Memczak, Henry A1 - Lauster, Daniel A1 - Kar, Parimal A1 - Di Lella, Santiago A1 - Volkmer, Rudolf A1 - Knecht, Volker A1 - Herrmann, Andreas A1 - Ehrentreich-Foerster, Eva A1 - Bier, Frank Fabian A1 - Stoecklein, Walter F. M. T1 - Anti-Hemagglutinin Antibody Derived Lead Peptides for Inhibitors of Influenza Virus Binding JF - PLoS one N2 - Antibodies against spike proteins of influenza are used as a tool for characterization of viruses and therapeutic approaches. However, development, production and quality control of antibodies is expensive and time consuming. To circumvent these difficulties, three peptides were derived from complementarity determining regions of an antibody heavy chain against influenza A spike glycoprotein. Their binding properties were studied experimentally, and by molecular dynamics simulations. Two peptide candidates showed binding to influenza A/Aichi/2/68 H3N2. One of them, termed PeB, with the highest affinity prevented binding to and infection of target cells in the micromolar region without any cytotoxic effect. PeB matches best the conserved receptor binding site of hemagglutinin. PeB bound also to other medical relevant influenza strains, such as human-pathogenic A/California/7/2009 H1N1, and avian-pathogenic A/MuteSwan/Rostock/R901/2006 H7N1. Strategies to improve the affinity and to adapt specificity are discussed and exemplified by a double amino acid substituted peptide, obtained by substitutional analysis. The peptides and their derivatives are of great potential for drug development as well as biosensing. Y1 - 2016 U6 - https://doi.org/10.1371/journal.pone.0159074 SN - 1932-6203 VL - 11 SP - 82 EP - 90 PB - PLoS CY - San Fransisco ER - TY - GEN A1 - Wessig, Pablo A1 - Bader, Denise A1 - Klier, Dennis Tobias A1 - Hettrich, Cornelia A1 - Bier, Frank Fabian T1 - Detecting carbohydrate–lectin interactions using a fluorescent probe based on DBD dyes N2 - Herein we present an efficient synthesis of a biomimetic probe with modular construction that can be specifically bound by the mannose binding FimH protein – a surface adhesion protein of E. coli bacteria. The synthesis combines the new and interesting DBD dye with the carbohydrate ligand mannose via a Click reaction. We demonstrate the binding to E. coli bacteria over a large concentration range and also present some special characteristics of those molecules that are of particular interest for the application as a biosensor. In particular, the mix-and-measure ability and the very good photo-stability should be highlighted here. T3 - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 314 KW - conformational-changes KW - green-i KW - protein KW - binding KW - assay Y1 - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-394382 SP - 1235 EP - 1238 ER - TY - JOUR A1 - Hüttl, Christine A1 - Hettrich, Cornelia A1 - Riedel, Melanie A1 - Henklein, Petra A1 - Rawel, Harshadrai Manilal A1 - Bier, Frank Fabian T1 - Development of Peptidyl Lysine Dendrons: 1,3-Dipolar Cycloaddition for Peptide Coupling and Antibody Recognition JF - Chemical biology & drug design N2 - A straightforward synthesis strategy to multimerize a peptide mimotopes for antibody B13-DE1 recognition is described based on lysine dendrons as multivalent scaffolds. Lysine dendrons that possess N-terminal alkyne residues at the periphery were quantitative functionalized with azido peptides using click chemistry. The solid-phase peptide synthesis (SPPS) allows preparing the peptide dendron in high purity and establishing the possibility of automation. The presented peptide dendron is a promising candidate as multivalent ligand and was used for antibody B13-DE1 recognition. The binding affinity increases with higher dendron generation without loss of specificity. The analysis of biospecific interaction between the synthesized peptide dendron and the antibody was done via surface plasmon resonance (SPR) technique. The presented results show a promising tool for investigations of antigen-antibody reactions. KW - click chemistry KW - lysine dendron KW - peptide mimotopes KW - solid-phase peptide synthesis KW - surface plasmon resonance Y1 - 2015 U6 - https://doi.org/10.1111/cbdd.12444 SN - 1747-0277 SN - 1747-0285 VL - 85 IS - 5 SP - 565 EP - 573 PB - Wiley-Blackwell CY - Hoboken ER - TY - JOUR A1 - Tanne, Johannes A1 - Jeoung, Jae-Hun A1 - Peng, Lei A1 - Yarman, Aysu A1 - Dietzel, Birgit A1 - Schulz, Burkhard A1 - Schad, Daniel A1 - Dobbek, Holger A1 - Wollenberger, Ursula A1 - Bier, Frank Fabian A1 - Scheller, Frieder W. T1 - Direct Electron Transfer and Bioelectrocatalysis by a Hexameric, Heme Protein at Nanostructured Electrodes JF - Electroanalysis : an international journal devoted to fundamental and practical aspects of electroanalysis N2 - A nanohybrid consisting of poly(3-aminobenzenesulfonic acid-co-aniline) and multiwalled carbon nanotubes [MWCNT-P(ABS-A)]) on a gold electrode was used to immobilize the hexameric tyrosine-coordinated heme protein (HTHP). The enzyme showed direct electron transfer between the heme group of the protein and the nanostructured surface. Desorption of the noncovalently bound heme from the protein could be excluded by control measurements with adsorbed hemin on aminohexanthiol-modified electrodes. The nanostructuring and the optimised charge characteristics resulted in a higher protein coverage as compared with MUA/MU modified electrodes. The adsorbed enzyme shows catalytic activity for the cathodic H2O2 reduction and oxidation of NADH. KW - HTHP KW - Nanohybrid KW - Poylaniline KW - Multiwalled carbon nanotube Y1 - 2015 U6 - https://doi.org/10.1002/elan.201500231 SN - 1040-0397 SN - 1521-4109 VL - 27 IS - 10 SP - 2262 EP - 2267 PB - Wiley-VCH CY - Weinheim ER - TY - JOUR A1 - Schmitz-Hertzberg, Sebastian-Tim A1 - Liese, Rick A1 - Terjung, Carsten A1 - Bier, Frank Fabian T1 - Towards a smart encapsulation system for small-sized electronic devices: a new approach JF - International journal of polymer science N2 - Miniaturized analytical chip devices like biosensors nowadays provide assistance in highly diverse fields of application such as point-of-care diagnostics and industrial bioprocess engineering. However, upon contact with fluids, the sensor requires a protective shell for its electrical components that simultaneously offers controlled access for the target analytes to the measuring units. We therefore developed a capsule that comprises a permeable and a sealed compartment consisting of variable polymers such as biocompatible and biodegradable polylactic acid (PLA) for medical applications or more economical polyvinyl chloride (PVC) and polystyrene (PS) polymers for bioengineering applications. Production of the sealed capsule compartments was performed by heat pressing of polymer pellets placed in individually designable molds. Controlled permeability of the opposite compartments was achieved by inclusion of NaCl inside the polymer matrix during heat pressing, followed by its subsequent release in aqueous solution. Correlating diffusion rates through the so made permeable capsule compartments were quantified for preselected model analytes: glucose, peroxidase, and polystyrene beads of three different diameters (1.4 mu m, 4.2 mu m, and 20.0 mu m). In summary, the presented capsule system turned out to provide sufficient shelter for small-sized electronic devices and gives insight into its potential permeability for defined substances of analytical interest. Y1 - 2014 U6 - https://doi.org/10.1155/2014/713603 SN - 1687-9422 SN - 1687-9430 PB - Hindawi Publishing Corp. CY - New York ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Yarman, Aysu A1 - Bachmann, Till A1 - Hirsch, Thomas A1 - Kubick, Stefan A1 - Renneberg, Reinhard A1 - Schumacher, Soeren A1 - Wollenberger, Ursula A1 - Teller, Carsten A1 - Bier, Frank Fabian ED - Gu, MB ED - Kim, HS T1 - Future of biosensors: a personal view JF - Advances in biochemical engineering, biotechnology JF - Advances in Biochemical Engineering-Biotechnology N2 - Biosensors representing the technological counterpart of living senses have found routine application in amperometric enzyme electrodes for decentralized blood glucose measurement, interaction analysis by surface plasmon resonance in drug development, and to some extent DNA chips for expression analysis and enzyme polymorphisms. These technologies have already reached a highly advanced level and need minor improvement at most. The dream of the "100-dollar' personal genome may come true in the next few years provided that the technological hurdles of nanopore technology or of polymerase-based single molecule sequencing can be overcome. Tailor-made recognition elements for biosensors including membrane-bound enzymes and receptors will be prepared by cell-free protein synthesis. As alternatives for biological recognition elements, molecularly imprinted polymers (MIPs) have been created. They have the potential to substitute antibodies in biosensors and biochips for the measurement of low-molecular-weight substances, proteins, viruses, and living cells. They are more stable than proteins and can be produced in large amounts by chemical synthesis. Integration of nanomaterials, especially of graphene, could lead to new miniaturized biosensors with high sensitivity and ultrafast response. In the future individual therapy will include genetic profiling of isoenzymes and polymorphic forms of drug-metabolizing enzymes especially of the cytochrome P450 family. For defining the pharmacokinetics including the clearance of a given genotype enzyme electrodes will be a useful tool. For decentralized online patient control or the integration into everyday "consumables' such as drinking water, foods, hygienic articles, clothing, or for control of air conditioners in buildings and cars and swimming pools, a new generation of "autonomous' biosensors will emerge. KW - Biosensors KW - Molecularly imprinted polymers KW - Personalized medicine Y1 - 2014 SN - 978-3-642-54143-8; 978-3-642-54142-1 U6 - https://doi.org/10.1007/10_2013_251 SN - 0724-6145 VL - 140 SP - 1 EP - 28 PB - Springer CY - Berlin ER - TY - JOUR A1 - Kersting, Sebastian A1 - Rausch, Valentina A1 - Bier, Frank Fabian A1 - von Nickisch-Rosenegk, Markus T1 - Rapid detection of Plasmodium falciparum with isothermal recombinase polymerase amplification and lateral flow analysis JF - Malaria journal N2 - Background: Nucleic acid amplification is the most sensitive and specific method to detect Plasmodium falciparum. However the polymerase chain reaction remains laboratory-based and has to be conducted by trained personnel. Furthermore, the power dependency for the thermocycling process and the costly equipment necessary for the read-out are difficult to cover in resource-limited settings. This study aims to develop and evaluate a combination of isothermal nucleic acid amplification and simple lateral flow dipstick detection of the malaria parasite for point-of-care testing. Methods: A specific fragment of the 18S rRNA gene of P. falciparum was amplified in 10 min at a constant 38 C using the isothermal recombinase polymerase amplification (RPA) method. With a unique probe system added to the reaction solution, the amplification product can be visualized on a simple lateral flow strip without further labelling. The combination of these methods was tested for sensitivity and specificity with various Plasmodium and other protozoa/bacterial strains, as well as with human DNA. Additional investigations were conducted to analyse the temperature optimum, reaction speed and robustness of this assay. Results: The lateral flow RPA (LF-RPA) assay exhibited a high sensitivity and specificity. Experiments confirmed a detection limit as low as 100 fg of genomic P. falciparum DNA, corresponding to a sensitivity of approximately four parasites per reaction. All investigated P. falciparum strains (n = 77) were positively tested while all of the total 11 non-Plasmodium samples, showed a negative test result. The enzymatic reaction can be conducted under a broad range of conditions from 30-45 degrees C with high inhibitory concentration of known PCR inhibitors. A time to result of 15 min from start of the reaction to read-out was determined. Conclusions: Combining the isothermal RPA and the lateral flow detection is an approach to improve molecular diagnostic for P. falciparum in resource-limited settings. The system requires none or only little instrumentation for the nucleic acid amplification reaction and the read-out is possible with the naked eye. Showing the same sensitivity and specificity as comparable diagnostic methods but simultaneously increasing reaction speed and dramatically reducing assay requirements, the method has potential to become a true point-of-care test for the malaria parasite. KW - Plasmodium falciparum KW - Recombinase polymerase amplification KW - RPA KW - PCR KW - Lateral flow KW - Point-of-care testing KW - Rapid test KW - Isothermal nucleic acid amplification Y1 - 2014 U6 - https://doi.org/10.1186/1475-2875-13-99 SN - 1475-2875 VL - 13 PB - BioMed Central CY - London ER - TY - JOUR A1 - Dechtrirat, Decha A1 - Gajovic-Eichelmann, Nenad A1 - Wojcik, Felix A1 - Hartmann, Laura A1 - Bier, Frank Fabian A1 - Scheller, Frieder W. T1 - Electrochemical displacement sensor based on ferrocene boronic acid tracer and immobilized glycan for saccharide binding proteins and E. coli JF - Biosensors and bioelectronics : the principal international journal devoted to research, design development and application of biosensors and bioelectronics N2 - Pathogens such as viruses and bacteria use their envelope proteins and their adhesin lectins to recognize the glycan residues presented on the cell surface of the target tissues. This principle of recognition is used in a new electrochemical displacement sensor for the protein concanavalin A (ConA). A gold electrode was first modified with a self-assembled monolayer of a thiolated mannose/OEG conjugate and a ferrocene boroxol derivative was pre-assembled as reporter molecule onto the mannose surface. The novel tracer molecule based on a 2-hydroxymethyl phenyl boronic acid derivative binds even at neutral pH to the saccharides which could expand the application towards biological samples (i.e., urine and feces). Upon the binding of ConA, the tracer was displaced and washed away from the sensor surface leading to a decrease in the electrochemical signal. Using square wave voltammetry (SWV), the concentration of ConA in the sample solution could be determined in the dynamic concentration range established from 38 nmol L-1 to 5.76 mu mol L-1 with a reproducible detection limit of 1 mu g mL(-1) (38 nmol L-1) based on the signal-to-noise ratio (S/N=3) with fast response of 15 min. The new reporter molecule showed a reduced non-specific displacement by BSA and ribonuclease A. The sensor was also successfully transferred to the first proof of principle for the detection of Escherichia coli exhibiting a detection limit of approximately 6 x 102 cells/mL Specificity of the displacement by target protein ConA and E. coli was demonstrated since the control proteins (i.e., BSA and RNaseA) and the control E. coli strain, which lack of type 1 fimbriae, were ineffective. (C) 2014 Elsevier B.V. All rights reserved. KW - Ferrocene benzoboroxol biosensor KW - Concanavalin A KW - Displacement KW - Escherichia coli KW - Ferrocene boronic acid KW - Self-assembled monolayer Y1 - 2014 U6 - https://doi.org/10.1016/j.bios.2014.02.028 SN - 0956-5663 SN - 1873-4235 VL - 58 SP - 1 EP - 8 PB - Elsevier CY - Oxford ER - TY - JOUR A1 - Kersting, Sebastian A1 - Rausch, Valentina A1 - Bier, Frank Fabian A1 - von Nickisch-Rosenegk, Markus T1 - Multiplex isothermal solid-phase recombinase polymerase amplification for the specific and fast DNA-based detection of three bacterial pathogens JF - Microchimica acta : analytical sciences based on micro- and nanomaterials N2 - We report on the development of an on-chip RPA (recombinase polymerase amplification) with simultaneous multiplex isothermal amplification and detection on a solid surface. The isothermal RPA was applied to amplify specific target sequences from the pathogens Neisseria gonorrhoeae, Salmonella enterica and methicillin-resistant Staphylococcus aureus (MRSA) using genomic DNA. Additionally, a positive plasmid control was established as an internal control. The four targets were amplified simultaneously in a quadruplex reaction. The amplicon is labeled during on-chip RPA by reverse oligonucleotide primers coupled to a fluorophore. Both amplification and spatially resolved signal generation take place on immobilized forward primers bount to expoxy-silanized glass surfaces in a pump-driven hybridization chamber. The combination of microarray technology and sensitive isothermal nucleic acid amplification at 38 degrees C allows for a multiparameter analysis on a rather small area. The on-chip RPA was characterized in terms of reaction time, sensitivity and inhibitory conditions. A successful enzymatic reaction is completed in < 20 min and results in detection limits of 10 colony-forming units for methicillin-resistant Staphylococcus aureus and Salmonella enterica and 100 colony-forming units for Neisseria gonorrhoeae. The results show this method to be useful with respect to point-of-care testing and to enable simplified and miniaturized nucleic acid-based diagnostics. KW - Isothermal amplification KW - RPA KW - Microchip KW - DNA sensor KW - Point-of-care Y1 - 2014 U6 - https://doi.org/10.1007/s00604-014-1198-5 SN - 0026-3672 SN - 1436-5073 VL - 181 IS - 13-14 SP - 1715 EP - 1723 PB - Springer CY - Wien ER - TY - JOUR A1 - Hoppe, Sebastian A1 - Bier, Frank Fabian A1 - von Nickisch-Rosenegk, Markus T1 - Identification of antigenic proteins of the nosocomial pathogen klebsiella pneumoniae JF - PLoS one N2 - The continuous expansion of nosocomial infections around the globe has become a precarious situation. Key challenges include mounting dissemination of multiple resistances to antibiotics, the easy transmission and the growing mortality rates of hospital-acquired bacterial diseases. Thus, new ways to rapidly detect these infections are vital. Consequently, researchers around the globe pursue innovative approaches for point-of-care devices. In many cases the specific interaction of an antigen and a corresponding antibody is pivotal. However, the knowledge about suitable antigens is lacking. The aim of this study was to identify novel antigens as specific diagnostic markers. Additionally, these proteins might be aptly used for the generation of vaccines to improve current treatment options. Hence, a cDNA-based expression library was constructed and screened via microarrays to detect novel antigens of Klebsiella pneumoniae, a prominent agent of nosocomial infections well-known for its extensive antibiotics resistance, especially by extended-spectrum beta-lactamases (ESBL). After screening 1536 clones, 14 previously unknown immunogenic proteins were identified. Subsequently, each protein was expressed in full-length and its immunodominant character examined by ELISA and microarray analyses. Consequently, six proteins were selected for epitope mapping and three thereof possessed linear epitopes. After specificity analysis, homology survey and 3d structural modelling, one epitope sequence GAVVALSTTFA of KPN_00363, an ion channel protein, was identified harboring specificity for K. pneumoniae. The remaining epitopes showed ambiguous results regarding the specificity for K. pneumoniae. The approach adopted herein has been successfully utilized to discover novel antigens of Campylobacter jejuni and Salmonella enterica antigens before. Now, we have transferred this knowledge to the key nosocomial agent, K. pneumoniae. By identifying several novel antigens and their linear epitope sites, we have paved the way for crucial future research and applications including the design of point-of-care devices, vaccine development and serological screenings for a highly relevant nosocomial pathogen. Y1 - 2014 U6 - https://doi.org/10.1371/journal.pone.0110703 SN - 1932-6203 VL - 9 IS - 10 PB - PLoS CY - San Fransisco ER - TY - JOUR A1 - Hovestaedt, Marc A1 - Memczak, Henry A1 - Pleiner, Dennis A1 - Zhang, Xin A1 - Rappich, Joerg A1 - Bier, Frank Fabian A1 - Stöcklein, Walter F. M. T1 - Characterization of a new maleimido functionalization of gold for surface plasmon resonance spectroscopy JF - Journal of molecular recognition : an international journal devoted to research on specific molecular recognition in chemistry, biology, biotechnology and medicine N2 - Para-maleimidophenyl (p-MP) modified gold surfaces have been prepared by one-step electrochemical deposition and used in surface plasmon resonance (SPR) studies. Therefore, a FITC mimotope peptide (MP1, 12 aa), a human mucin 1 epitope peptide (MUC, 9 aa) and a protein with their specific antibodies were used as model systems. The peptides were modified with an N-terminal cysteine for covalent and directed coupling to the maleimido functionalized surface by means of Michael addition. The coupling yield of the peptide, the binding characteristics of antibody and the unspecific adsorption of the analytes were investigated. The results expand the spectrum of biosensors usable with p-MP by widely used SPR and support its potential to be versatile for several electrochemical and optical biosensors. This allows the combination of an electrochemical and optical read-out for a broad variety of biomolecular interactions on the same chip. Copyright (c) 2014 John Wiley & Sons, Ltd. KW - biosensor KW - surface plasmon resonance KW - diazonium coupling KW - maleimidophenyl KW - cys-peptide KW - aryl diazonium salts Y1 - 2014 U6 - https://doi.org/10.1002/jmr.2396 SN - 0952-3499 SN - 1099-1352 VL - 27 IS - 12 SP - 707 EP - 713 PB - Wiley-Blackwell CY - Hoboken ER - TY - JOUR A1 - Stanke, Sandra A1 - Bier, Frank Fabian A1 - Hoezel, Ralph T1 - Fluid streaming above interdigitated electrodes in dielectrophoresis experiments JF - Electrophoresis : microfluidics, nanoanalysis & proteomics N2 - For the investigation of alternating current electrokinetic effects, a system is presented that allows for the simultaneous observation of fluid flow above and around microelectrodes in all three directions in space. Beside the usual microscopical view from top, lateral observation through the same objective is made possible by two small mirrors that are placed next to the electrodes. Fluid flow and movement of fluorescent nanoparticles above interdigitated electrodes are monitored by fluorescence microscopy and digital imaging and are further analysed by image processing. Field frequencies are varied from 10 Hz to 1 GHz at up to 10V(rms). Electrical conductivity of the fluid is monitored in situ in the actual measuring chamber. KW - Dielectrophoresis KW - Fluid streaming KW - Nanobead Y1 - 2011 U6 - https://doi.org/10.1002/elps.201100096 SN - 0173-0835 VL - 32 IS - 18 SP - 2448 EP - 2455 PB - Wiley-Blackwell CY - Malden ER - TY - JOUR A1 - Breitenstein, Michael A1 - Nielsen, Peter E. A1 - Hölzel, Ralph A1 - Bier, Frank Fabian T1 - DNA-nanostructure-assembly by sequential spotting JF - Journal of nanobiotechnology N2 - Background: The ability to create nanostructures with biomolecules is one of the key elements in nanobiotechnology. One of the problems is the expensive and mostly custom made equipment which is needed for their development. We intended to reduce material costs and aimed at miniaturization of the necessary tools that are essential for nanofabrication. Thus we combined the capabilities of molecular ink lithography with DNA-self-assembling capabilities to arrange DNA in an independent array which allows addressing molecules in nanoscale dimensions. Results: For the construction of DNA based nanostructures a method is presented that allows an arrangement of DNA strands in such a way that they can form a grid that only depends on the spotted pattern of the anchor molecules. An atomic force microscope (AFM) has been used for molecular ink lithography to generate small spots. The sequential spotting process allows the immobilization of several different functional biomolecules with a single AFM-tip. This grid which delivers specific addresses for the prepared DNA-strand serves as a two-dimensional anchor to arrange the sequence according to the pattern. Once the DNA-nanoarray has been formed, it can be functionalized by PNA (peptide nucleic acid) to incorporate advanced structures. Conclusions: The production of DNA-nanoarrays is a promising task for nanobiotechnology. The described method allows convenient and low cost preparation of nanoarrays. PNA can be used for complex functionalization purposes as well as a structural element. Y1 - 2011 U6 - https://doi.org/10.1186/1477-3155-9-54 SN - 1477-3155 VL - 9 IS - 11 PB - BioMed Central CY - London ER - TY - INPR A1 - Baret, Jean-Christophe A1 - Belder, Detlev A1 - Bier, Frank Fabian A1 - Cao, Jialan A1 - Gruschke, Oliver A1 - Hardt, Steffen A1 - Kirschbaum, Michael A1 - Koehler, J. Michael A1 - Schumacher, Soeren A1 - Urban, G. A. A1 - Viefhues, Martina T1 - Contributors to the 10th Anniversary Germany issue T2 - LAB on a chip : miniaturisation for chemistry and biology Y1 - 2012 U6 - https://doi.org/10.1039/c1lc90139g SN - 1473-0197 VL - 12 IS - 3 SP - 419 EP - 421 PB - Royal Society of Chemistry CY - Cambridge ER - TY - JOUR A1 - Hoppe, Sebastian A1 - Bier, Frank Fabian A1 - von Nickisch-Rosenegk, Markus T1 - Microarray-based method for screening of immunogenic proteins from bacteria JF - Journal of nanobiotechnology N2 - Background: Detection of immunogenic proteins remains an important task for life sciences as it nourishes the understanding of pathogenicity, illuminates new potential vaccine candidates and broadens the spectrum of biomarkers applicable in diagnostic tools. Traditionally, immunoscreenings of expression libraries via polyclonal sera on nitrocellulose membranes or screenings of whole proteome lysates in 2-D gel electrophoresis are performed. However, these methods feature some rather inconvenient disadvantages. Screening of expression libraries to expose novel antigens from bacteria often lead to an abundance of false positive signals owing to the high cross reactivity of polyclonal antibodies towards the proteins of the expression host. A method is presented that overcomes many disadvantages of the old procedures. Results: Four proteins that have previously been described as immunogenic have successfully been assessed immunogenic abilities with our method. One protein with no known immunogenic behaviour before suggested potential immunogenicity. We incorporated a fusion tag prior to our genes of interest and attached the expressed fusion proteins covalently on microarrays. This enhances the specific binding of the proteins compared to nitrocellulose. Thus, it helps to reduce the number of false positives significantly. It enables us to screen for immunogenic proteins in a shorter time, with more samples and statistical reliability. We validated our method by employing several known genes from Campylobacter jejuni NCTC 11168. Conclusions: The method presented offers a new approach for screening of bacterial expression libraries to illuminate novel proteins with immunogenic features. It could provide a powerful and attractive alternative to existing methods and help to detect and identify vaccine candidates, biomarkers and potential virulence-associated factors with immunogenic behaviour furthering the knowledge of virulence and pathogenicity of studied bacteria.zeige weniger Y1 - 2012 U6 - https://doi.org/10.1186/1477-3155-10-12 SN - 1477-3155 VL - 10 PB - BioMed Central CY - London ER - TY - JOUR A1 - Linck, Lena A1 - Reiss, Edda A1 - Bier, Frank Fabian A1 - Resch-Genger, Ute T1 - Direct labeling rolling circle amplification as a straightforward signal amplification technique for biodetection formats JF - Analytical methods : advancing methods and applications N2 - Biodetection formats, such as DNA and antibody microarrays, are valuable tools in the life sciences, but for some applications, the detection limits are insufficient. A straightforward strategy to obtain signal amplification is the rolling circle amplification (RCA), an easy, isothermal, and enzymatic nucleic acid synthesis that has already been employed successfully to increase the signal yield for several single-analyte and multiplexing assays in conjunction with hybridization probes. Here, we systematically investigated the parameters responsible for the RCA driven signal amplification with fluorescent labels, such as the type of fluorophore chosen, labeling strategy, composition of reaction solution, and number of handling steps. In labeling strategies, post-synthetic labeling via a Cy3-hybridization probe was compared to the direct incorporation of fluorescent Cy3-dUTP and DY-555-dUTP into the nascent strand during synthesis. With our direct labeling protocol, the assay's runtime and handling steps could be reduced while the signal yield was increased. These features are very attractive for many detection formats but especially for point-of-care diagnostic kits that need to be simple enough to be performed by scientifically untrained personnel. Y1 - 2012 U6 - https://doi.org/10.1039/c2ay05760c SN - 1759-9660 VL - 4 IS - 5 SP - 1215 EP - 1220 PB - Royal Society of Chemistry CY - Cambridge ER - TY - JOUR A1 - Stech, Marlitt A1 - Merk, Helmut A1 - Schenk, Jörg A. A1 - Stöcklein, Walter F. M. A1 - Wüstenhagen, Doreen Anja A1 - Micheel, Burkhard A1 - Duschl, Claus A1 - Bier, Frank Fabian A1 - Kubick, Stefan T1 - Production of functional antibody fragments in a vesicle-based eukaryotic cell-free translation system JF - Journal of biotechnology N2 - Cell-free protein synthesis is of increasing interest for the rapid and high-throughput synthesis of many proteins, in particular also antibody fragments. In this study, we present a novel strategy for the production of single chain antibody fragments (scFv) in a eukaryotic in vitro translation system. This strategy comprises the cell-free expression, isolation and label-free interaction analysis of a model antibody fragment synthesized in two differently prepared insect cell lysates. These lysates contain translocationally active microsomal structures derived from the endoplasmic reticulum (ER), allowing for posttranslational modifications of cell-free synthesized proteins. Both types of these insect cell lysates enable the synthesis and translocation of scFv into ER-derived vesicles. However, only the one that has a specifically adapted redox potential yields functional active antibody fragments. We have developed a new methodology for the isolation of functional target proteins based on the translocation of cell-free produced scFv into microsomal structures and subsequent collection of protein-enriched vesicles. Antibody fragments that have been released from these vesicles are shown to be well suited for label-free binding studies. Altogether, these results show the potential of insect cell lysates for the production, purification and selection of antibody fragments in an easy-to-handle and time-saving manner. KW - Cell-free KW - In vitro translation KW - Single chain antibody (scFv) KW - Insect lysate KW - Surface plasmon resonance Y1 - 2012 U6 - https://doi.org/10.1016/j.jbiotec.2012.08.020 SN - 0168-1656 VL - 164 IS - 2 SP - 220 EP - 231 PB - Elsevier CY - Amsterdam ER - TY - JOUR A1 - Sachse, Rita A1 - Wüstenhagen, Doreen Anja A1 - Samalikova, Maria A1 - Gerrits, Michael A1 - Bier, Frank Fabian A1 - Kubick, Stefan T1 - Synthesis of membrane proteins in eukaryotic cell-free systems JF - Engineering in life sciences : Industry, Environment, Plant, Food N2 - Cell-free protein synthesis (CFPS) is a valuable method for the fast expression of difficult-to-express proteins as well as posttranslationally modified proteins. Since cell-free systems circumvent possible cytotoxic effects caused by protein overexpression in living cells, they significantly enlarge the scale and variety of proteins that can be characterized. We demonstrate the high potential of eukaryotic CFPS to express various types of membrane proteins covering a broad range of structurally and functionally diverse proteins. Our eukaryotic cell-free translation systems are capable to provide high molecular weight membrane proteins, fluorescent-labeled membrane proteins, as well as posttranslationally modified proteins for further downstream analysis. KW - Cell-free protein expression KW - In vitro protein synthesis KW - Labeled membrane proteins KW - Synthetic glycoprotein Y1 - 2013 U6 - https://doi.org/10.1002/elsc.201100235 SN - 1618-0240 VL - 13 IS - 1 SP - 39 EP - 48 PB - Wiley-Blackwell CY - Hoboken ER - TY - CHAP A1 - Memczak, Henry A1 - Lauster, Daniel A1 - Herrmann, Andreas A1 - Stöcklein, Walter F. M. A1 - Bier, Frank Fabian T1 - Novel hemagglutinin-binding peptides for biosensing and inhibition of Influenza Viruses T2 - Biopolymers Y1 - 2013 SN - 0006-3525 SN - 1097-0282 VL - 100 IS - 3 SP - 255 EP - 255 PB - Wiley-Blackwell CY - Hoboken ER - TY - JOUR A1 - Hoppe, Sebastian A1 - Bier, Frank Fabian A1 - von Nickisch-Rosenegk, Markus T1 - Rapid identification of novel immunodominant proteins and characterization of a specific linear epitope of campylobacter jejuni JF - PLoS one N2 - Campylobacter jejuni remains one of the major gut pathogens of our time. Its zoonotic nature and wide-spread distribution in industrialized countries calls for a quick and reliable diagnostic tool. Antibody-based detection presents a suitable means to identify pathogenic bacteria. However, the knowledge about immunodominant targets is limited. Thus, an approach is presented, which allows for the rapid screening of numerous cDNA derived expression clones to identify novel antigens. The deeper understanding of immunodominant proteins assists in the design of diagnostic tools and furthers the insight into the bacterium's pathogenicity as well as revealing potential candidates for vaccination. We have successfully screened 1536 clones of an expression library to identify 22 proteins that have not been described as immunodominant before. After subcloning the corresponding 22 genes and expression of full-length proteins, we investigated the immunodominant character by microarrays and ELISA. Subsequently, seven proteins were selected for epitope mapping. For cj0669 and cj0920c linear epitopes were identified. For cj0669, specificity assays revealed a specific linear epitope site. Consequently, an eleven amino acid residue sequence TLIKELKRLGI was analyzed via alanine scan, which revealed the glycine residue to be significant for binding of the antibody. The innovative approach presented herein of generating cDNAs of prokaryotes in combination with a microarray platform rendering time-consuming purification steps obsolete has helped to illuminate novel immunodominant proteins of C. jejuni. The findings of a specific linear epitope pave the way for a plethora of future research and the potential use in diagnostic applications such as serological screenings. Moreover, the current approach is easily adaptable to other highly relevant bacteria making it a formidable tool for the future discovery of antigens and potential biomarkers. Consequently, it is desirable to simplify the identification of structural epitopes, as this would extend the spectrum of novel epitopes to be detected. Y1 - 2013 U6 - https://doi.org/10.1371/journal.pone.0065837 SN - 1932-6203 VL - 8 IS - 5 PB - PLoS CY - San Fransisco ER - TY - JOUR A1 - Hüttl, Christine A1 - Hettrich, Cornelia A1 - Miller, Reinhard A1 - Paulke, Bernd-Reiner A1 - Henklein, Petra A1 - Rawel, Harshadrai Manilal A1 - Bier, Frank Fabian T1 - Self-assembled peptide amphiphiles function as multivalent binder with increased hemagglutinin affinity JF - BMC biotechnology N2 - Background: A promising way in diagnostic and therapeutic applications is the development of peptide amphiphiles (PAs). Peptides with a palmitic acid alkylchain were designed and characterized to study the effect of the structure modifications on self-assembling capabilities and the multiple binding capacity to hemagglutinin (HA), the surface protein of influenza virus type A. The peptide amphiphiles consists of a hydrophilic headgroup with a biological functionality of the peptide sequence and a chemically conjugated hydrophobic tail. In solution they self-assemble easily to micelles with a hydrophobic core surrounded by a closely packed peptide-shell. Results: In this study the effect of a multiple peptide binding partner to the receptor binding site of HA could be determined with surface plasmon resonance measurements. The applied modification of the peptides causes signal amplification in relationship to the unmodified peptide wherein the high constant specificity persists. The molecular assembly of the peptides was characterized by the determination of critical micelle concentration (CMC) with concentration of 10(-5) M and the colloidal size distribution. Conclusion: The modification of the physico-chemical parameters by producing peptide amphiphiles form monomeric structures which enhances the binding affinity and allows a better examination of the interaction with the virus surface protein hemagglutinin. KW - CMC KW - Influenza virus detection KW - Micelle KW - PAs KW - Surface plasmon resonance Y1 - 2013 U6 - https://doi.org/10.1186/1472-6750-13-51 SN - 1472-6750 VL - 13 IS - 22 PB - BioMed Central CY - London ER - TY - JOUR A1 - Schmitz-Hertzberg, Sebastian-Tim A1 - Mak, Wing Cheung A1 - Lai, Kwok Kei A1 - Teller, Carsten A1 - Bier, Frank Fabian T1 - Multifactorial design of Poly(D, L-lactic-co-glycolic acid) capsules with various release properties for differently sized filling agents JF - Journal of applied polymer science N2 - The hydrolytic degradation and corresponding content release of capsules made of poly(d,l-lactic-co-glycolic acid) (PLGA) strongly depends on the composition and material properties of the initially applied copolymer. Consecutive or simultaneous release from capsule batches of combinable material compositions, therefore, offers high control over the bioavailability of an encapsulated drug. The keynote of this study was the creation of a superordinated database that addressed the correlation between the release kinetics of filling agents with different molecular weights from PLGA capsules of alternating composition. Fluorescein isothiocyanate (FITC)-dextran (with molecular weights of 4, 40, and 2000 kDa) was chosen as a model analyte, whereas the copolymers were taken from various 50:50 PLGA, 75:25 PLGA, and polylactide blends. With reference to recent publications, the capsule properties, such as the size, morphology, and encapsulation efficiency, were further modified during production. Hence, uniform microdisperse and polydisperse submicrometer nanocapsules were prepared by two different water-in-oil-in-water emulsification techniques, and additional effects on the size and morphology were achieved by capsule solidification in two different sodium salt buffers. The qualitative and quantitative examination of the physical capsule properties was performed by confocal laser scanning microscopy, scanning electron microscopy, and Coulter counting techniques to evaluate the capsule size distribution and the morphological appearance of the different batches. The corresponding agent release was quantified by fluorescence measurement of the FITC-dextran in the incubation media and by the direct measurement of the capsule brightness via fluorescence microscopy. In summary, the observed agent release showed a highly controllable flexibility depending on the PLGA blends, preparation methods, and molecular weight of the used filling substances. (c) 2013 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 130: 4219-4228, 2013 KW - biodegradable copolymers (PLGA) KW - microcapsules KW - submicrometer KW - nanocapsules KW - FITC-dextran release KW - drug delivery system KW - biomedical applications Y1 - 2013 U6 - https://doi.org/10.1002/app.39537 SN - 0021-8995 SN - 1097-4628 VL - 130 IS - 6 SP - 4219 EP - 4228 PB - Wiley-Blackwell CY - Hoboken ER - TY - JOUR A1 - Grießner, Matthias A1 - Hartig, Dave A1 - Christmann, Alexander A1 - Ehrentreich-Förster, Eva A1 - Warsinke, Axel A1 - Bier, Frank Fabian T1 - Surface regeneration of microfluidic microarray printheads through plasma techniques N2 - This work describes a method for surface regeneration of microfluidic microarray printheads through plasma techniques. Modification procedures were chosen in a way to obtain high reproducibility with a minimum of time consumption. The idea behind this is a complete regeneration of a microarray printhead before or after usage to achieve best printing results over a typical print job. A sequence of low-pressure oxygen-plasma and plasma polymerization with hexamethyldisiloxane (HMDSO) was used to regenerate printheads. Proof of the concept is given through quality control performed with a spotter implemented CCD camera, contact angle measurements and a typical hybridization experiment. Stable printing results were obtained over 3000 activations showing that the presented method is suitable for treatment of microarray printheads. Y1 - 2010 UR - http://iopscience.iop.org/0960-1317/ U6 - https://doi.org/10.1088/0960-1317/20/3/037002 SN - 0960-1317 ER - TY - JOUR A1 - Reiss, Edda A1 - Hoelzel, Ralph A1 - Bier, Frank Fabian T1 - Synthesis and stretching of rolling circle amplification products in a flow-through system N2 - Enzymatic isothermal rolling circle amplification (RCA) produces long concatemeric single-stranded DNA (ssDNA) molecules if a small circular ssDNA molecule is applied as the template. A method is presented here in which the RCA reaction is carried out in a flow-through system, starting from isolated surface-tethered DNA primers. This approach combines gentle fluidic handling of the single-stranded RCA products, such as staining or stretching via a receding meniscus, with the option of simultaneous (fluorescence) microscopic observation. It is shown that the stretched and surface-attached RCA products are accessible for hybridization of complementary oligonucleotides, which demonstrates their addressability by complementary base pairing. The long RCA products should be well suited to bridge the gap between biomolecular nanoscale building-blocks and structures at the micro- and macroscale, especially at the single- molecule level presented here. Y1 - 2009 UR - http://www3.interscience.wiley.com/cgi-bin/jhome/107640323 U6 - https://doi.org/10.1002/smll.200900319 SN - 1613-6810 ER - TY - JOUR A1 - Grießner, Matthias A1 - Broeker, Patrick A1 - Lehmann, André A1 - Ehrentreich-Förster, Eva A1 - Bier, Frank Fabian T1 - Detection of angiotensin II type 1 receptor ligands by a cell-based assay N2 - This work describes a cell-based assay that does not depend on radioactivity or laboratory animals for the detection of ligands of angiotensin II type 1 receptor (AT(1)R). The assay makes use of stable transfected Chinese hamster ovary cells (CHO-AT(1)R) expressing the AT(1)R. A sequential saturation assay principle was used in which receptor binding sites of the CHO-AT(1)R cells are blocked by the analyte in a concentration-dependent manner. Afterwards, TAMRA-angiotensin II, a fluorescence-labeled ligand, was added to bind to the remaining free binding sites of the receptor. In consequence, the fluorescence signal determined is inversely proportional to the concentration of the analyte. Y1 - 2009 UR - http://www.springerlink.com/content/100417 U6 - https://doi.org/10.1007/s00216-009-3074-4 SN - 1618-2642 ER - TY - JOUR A1 - Andresen, Dennie A1 - von Nickisch-Rosenegk, Markus A1 - Bier, Frank Fabian T1 - Helicase-dependent amplification : use in OnChip amplification and potential for point-of-care diagnostics N2 - Isothermal amplification technologies are emerging on the horizon that could have the potential to pose as alternatives to PCR in terms of sensitivity and ease of use. One of the most recent isothermal technologies is helicase- dependent amplification (HDA). This technology uses the helicase's capability to disrupt the hydrogen bonds of a Watson-Crick base pair in order to separate dsDNA. A denaturation step, as is used in PCR, is no longer required. This gives rise to new, less expensive and less complicated designs for point-of-care devices and 'Lab on Chip' systems. Helicase-dependent OnChip-amplification (OnChip-HDA) is a further step into this direction as it integrates the HDA technology with microarray technology and its power of multiplexing. This special report will give an overview on the HDA and OnChip-HDA technology, and its potential for point-of-care diagnostics. Y1 - 2009 UR - http://www.expert-reviews.com/loi/erm U6 - https://doi.org/10.1586/erm.09.46 SN - 1473-7159 ER - TY - JOUR A1 - Andresen, Dennie A1 - von Nickisch-Rosenegk, Markus A1 - Bier, Frank Fabian T1 - Helicase dependent OnChip-amplification and its use in multiplex pathogen detection N2 - Background: The need for fast, specific and sensitive multiparametric detection methods is an ever growing demand in molecular diagnostics. Here we report on a newly developed method, the helicase dependent Onchip amplification (OnChip-HDA). This approach integrates the analysis and detection in one single reaction thus leading to time and cost savings in multiparametric analysis. Methods: HDA is an isothermal amplification method that is not depending on thermocycling as known from PCR due to the helicases' ability to unwind DNA double-strands. We have combined the HDA with microarray based detection, making it suitable for multiplex detection. As an example we used the Onchip HDA in single and multiplex amplifications for the detection of the two pathogens N. gonorrhoeae and S. aureus directly on surface bound primers. Results: We have successfully shown the OnChip-HDA and applied it for single- and duplex- detection of the pathogens N. gonorrhoeae and S. aureus. Conclusion: We have developed a new method, the OnChip-HDA for the multiplex detection of pathogens. Its simplicity in reaction setup and potential for miniaturization and multiparametric analysis is advantageous for the integration in miniaturized Lab on Chip systems, e.g. needed in point of care diagnostics. Y1 - 2009 UR - http://www.sciencedirect.com/science/journal/00098981 U6 - https://doi.org/10.1016/j.cca.2009.03.021 SN - 0009-8981 ER - TY - JOUR A1 - Andresen, Heiko A1 - Grotzinger, Carsten A1 - Zarse, Kim A1 - Kreuzer, Oliver Johannes A1 - Ehrentreich-Förster, Eva A1 - Bier, Frank Fabian T1 - Functional peptide microarrays for specific and sensitive antibody diagnostics N2 - Peptide microarrays displaying biologically active small synthetic peptides in a high-density format provide an attractive technology to probe complex samples for the presence and/or function of protein analytes. We present a new approach for manufacturing functional peptide microarrays for molecular immune diagnostics. Our method relies on the efficiency of site-specific solution-phase coupling of biotinylated synthetic peptides to NeutrAvidin (NA) and localized microdispensing of peptide-NA-complexes onto activated glass surfaces. Antibodies are captured in a sandwich manner between surface immobilized peptide probes and fluorescence-labeled secondary antibodies. Our work includes a total of 54 peptides derived from immunodominant linear epitopes of the T7 phage capsid protein, Herpes simplex virus glycoprotein D, c-myc protein, and three domains of the Human coronavirus polymerase polyprotein and their cognate mAbs. By using spacer molecules of different type and length for NA-mediated peptide presentation, we show that the incorporation of a minimum spacer length is imperative for antibody binding, whereas the peptide immobilization direction has only secondary importance for antibody affinity and binding. We further demonstrate that the peptide array is capable of detecting low-picomolar concentrations of mAbs in buffered solutions and diluted human serum with high specificity Y1 - 2006 UR - http://www3.interscience.wiley.com/cgi-bin/jhome/76510741 U6 - https://doi.org/10.1002/pmic.200500343 SN - 1615-9853 ER - TY - JOUR A1 - Andresen, Heiko A1 - Grötzinger, Carsten A1 - Zarse, Kim A1 - Birringer, Marc A1 - Hessenius, Carsten A1 - Kreuzer, Oliver Johannes A1 - Ehrentreich-Förster, Eva A1 - Bier, Frank Fabian T1 - Peptide microarrays with site-specifically immobilized synthetic peptides for antibody diagnostics N2 - Peptide microarrays bear the potential to discover molecular recognition events on protein level, particularly in the field of molecular immunology, in a manner and with an efficiency comparable to the performance of DNA microarrays. We developed a novel peptide microarray platform for the detection of antibodies in liquid samples. The system comprises site-specific solution phase coupling of biotinylated peptides to NeutrAvidin, localized microdispensing of peptide-NeutrAvidin conjugates onto activated glass slides and a fluorescence immuno sandwich assay format for antibody capture and detection. Our work includes synthetic peptides deduced from amino acid sequences of immunodominant linear epitopes, such as the T7 phage capsid protein, Herpes simplex virus glycoprotein D, c-myc protein and three domains of the Human coronavirus 229E polymerase polyprotein. We demonstrate that our method produces peptide arrays with excellent spot morphology which are capable of specific and sensitive detection of monoclonal antibodies from fluid samples. Y1 - 2006 UR - http://www.sciencedirect.com/science/journal/09254005 U6 - https://doi.org/10.1016/j.snb.2005.07.033 SN - 0925-4005 ER - TY - JOUR A1 - Bier, Frank Fabian A1 - Scheller, Frieder W. A1 - Klingbeil, Mandy A1 - Oßwald, U. T1 - Biosensoren und Teststreifen für die Umwelt- und Lebensmittelanalytik : eine Übersicht Y1 - 1993 ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Bier, Frank Fabian A1 - Neumann, B. T1 - Bioindikation in aquatischen Ökosystemen : Bioindikation in limnischen und küstennahen Ökosystemen ; Grundlagen, Verfahren und Methoden Y1 - 1994 PB - Fischer CY - Jena ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Makower, Alexander A1 - Ghindilis, A. L. A1 - Bier, Frank Fabian A1 - Ehrentreich-Förster, Eva A1 - Wollenberger, Ursula A1 - Bauer, Christian G. A1 - Micheel, Burkhard A1 - Pfeiffer, Dorothea A1 - Szeponik, Jan A1 - Michael, N. A1 - Kaden, H. T1 - Enzyme sensors for subnanomolar concentrations Y1 - 1995 ER - TY - JOUR A1 - Jin, Wen A1 - Wollenberger, Ursula A1 - Bier, Frank Fabian A1 - Scheller, Frieder W. T1 - Construction and characterization of multi-layer-enzyme electrode : covalent binding of quinoprotein glucose dehydrogenase onto gold electrodes Y1 - 1995 ER - TY - JOUR A1 - Ghindilis, A. L. A1 - Makower, Alexander A1 - Bauer, Christian G. A1 - Bier, Frank Fabian A1 - Scheller, Frieder W. T1 - Determination of p-aminophenol and catecholamines at picomolar concentrations based on recycling enzyme amplification Y1 - 1995 ER - TY - JOUR A1 - Song, Min Ik A1 - Bier, Frank Fabian A1 - Scheller, Frieder W. T1 - A method to detect superoxide radicals using teflon membrane and superoxide dismutase Y1 - 1995 ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Schubert, Florian A1 - Bier, Frank Fabian T1 - Vom Biosensor zur Nanobiotechnologie Y1 - 1995 ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Bier, Frank Fabian A1 - Pfeiffer, Dorothea T1 - Biosensoren : Grundlagen und Anwendungen Y1 - 1995 ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Makower, Alexander A1 - Bier, Frank Fabian A1 - Wollenberger, Ursula A1 - Ghindilis, A. L. A1 - Eremenko, A. V. A1 - Pfeiffer, Dorothea T1 - Enzymsensoren zur Bestimmung subnanomolarer Konzentrationen Y1 - 1995 ER - TY - JOUR A1 - Bauer, Christian G. A1 - Eremenko, A. V. A1 - Ehrentreich-Förster, Eva A1 - Bier, Frank Fabian A1 - Makower, Alexander A1 - Halsall, H. B. A1 - Heineman, W. R. A1 - Scheller, Frieder W. T1 - Zeptomole-detecting biosensor for alkaline phosphatase in an electroche mical immunoassay for 2,4- dichlorophenoacetic acid Y1 - 1996 ER - TY - JOUR A1 - Bier, Frank Fabian A1 - Ehrentreich-Förster, Eva A1 - Bauer, Christian G. A1 - Scheller, Frieder W. T1 - High sensitive competitive immunodetection of 2,4-dichlorophenoxyacetic acid using enzymatic amplification with electrochemical detection Y1 - 1996 ER - TY - JOUR A1 - Bier, Frank Fabian A1 - Ehrentreich-Förster, Eva A1 - Scheller, Frieder W. A1 - Makower, Alexander A1 - Eremenko, A. V. A1 - Wollenberger, Ursula A1 - Bauer, Christian G. A1 - Pfeiffer, Dorothea A1 - Micheel, Burkhard T1 - Ultrasensitive biosensors Y1 - 1996 ER - TY - JOUR A1 - Bier, Frank Fabian A1 - Ehrentreich-Förster, Eva A1 - Makower, Alexander A1 - Scheller, Frieder W. T1 - An enzymatic amplification cycle for high sensitive immunoassay Y1 - 1996 ER - TY - JOUR A1 - Bier, Frank Fabian A1 - Scheller, Frieder W. T1 - Label-free observation of DNA-hybridisation and endonuclease activity on a wave guide surface using a grating coupler Y1 - 1996 ER - TY - JOUR A1 - Bier, Frank Fabian A1 - Ehrentreich-Förster, Eva A1 - Scheller, Frieder W. T1 - Amplifying bienzyme cycle-linked immunoassays for determination of 2,4- dichlorphenoxyacetic acid Y1 - 1996 ER - TY - JOUR A1 - Jin, Wen A1 - Wollenberger, Ursula A1 - Bier, Frank Fabian A1 - Makower, Alexander A1 - Schiller, Frieder W. T1 - Electron transfer between cytochrome c and copper enzymes Y1 - 1996 ER - TY - JOUR A1 - Bier, Frank Fabian A1 - Fürste, J. P. T1 - Nucleic acid based sensors Y1 - 1997 ER - TY - JOUR A1 - Bier, Frank Fabian A1 - Ehrentreich-Förster, Eva A1 - Dölling, R. A1 - Eremenko, A. V. A1 - Scheller, Frieder W. T1 - A redox-label immunosensor on basis of a bi-enzyme electrode Y1 - 1997 ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Bier, Frank Fabian A1 - Gajovic, Nenad T1 - Biosensoren und Teststreifen für die Umwelt- und Lebensmittelanalytik Y1 - 1997 ER - TY - JOUR A1 - Stöcklein, Walter F. M. A1 - Makower, Alexander A1 - Bier, Frank Fabian A1 - Scheller, Frieder W. T1 - Enzyme sensors and enzyme amplifification systems Y1 - 1997 ER - TY - JOUR A1 - Makower, Alexander A1 - Barmin, Anatoli V. A1 - Morzunova, T. A1 - Eremenko, Arkadi V. A1 - Bier, Frank Fabian A1 - Scheller, Frieder W. T1 - Affinity enzymomoetric assay for detection of organophosphorus compounds Y1 - 1997 ER - TY - JOUR A1 - Bier, Frank Fabian A1 - Fürste, J. P. A1 - Kleinjung, Frank A1 - Erdmann, V. A. A1 - Scheller, Frieder W. T1 - Nukleinsäuren als Basis für Biosensoren Y1 - 1997 ER - TY - JOUR A1 - Bier, Frank Fabian A1 - Kleinjung, Frank A1 - Scheller, Frieder W. T1 - Real time measurement of nucleic acid hybridization using evanescent wave sensors - step towards the genosensor Y1 - 1997 ER - TY - THES A1 - Bier, Frank Fabian T1 - Biomolekulare Erkennung und Signaltransduktion in Affinitätssensoren Y1 - 1997 CY - Potsdam ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Kleinjung, Frank A1 - Bier, Frank Fabian A1 - Markower, Alexander A1 - Neumann, Barbara A1 - Wollenberger, Ursula A1 - Kurochkin, Iliya N. A1 - Eremenko, Arkadi V. A1 - Barmin, Anatoli V. A1 - Klußmann, Sven A1 - Fürste, Jens-Peter A1 - Erdmann, Volker A. A1 - Mansuy, D. T1 - New recognition elements in biosensing Y1 - 1998 ER - TY - JOUR A1 - Kleinjung, Frank A1 - Klußmann, S. A1 - Erdmann, V. A. A1 - Scheller, Frieder W. A1 - Fürste, J. P. A1 - Bier, Frank Fabian T1 - Novel binders in biosensorics : hight affinity RNA for smal analytes Y1 - 1998 ER - TY - JOUR A1 - Schmidt, Peter Michael A1 - Matthes, E. A1 - Scheller, Frieder W. A1 - Bier, Frank Fabian T1 - Nachweis der Telomeraseaktivität in Zellkulturen mittels eines faseroptischen Sensors Y1 - 2001 ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Bauer, Christian G. A1 - Markower, Alexander A1 - Wollenberger, Ursula A1 - Warsinke, Axel A1 - Bier, Frank Fabian T1 - Coupling of immunoassays with enzymatic recycling electrodes Y1 - 2001 ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Bier, Frank Fabian T1 - Trends in der Bioanalytik Y1 - 2002 ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Bauer, Christian G. A1 - Makower, Alexander A1 - Wollenberger, Ursula A1 - Warsinke, Axel A1 - Bier, Frank Fabian T1 - Immunoassays using enzymatic amplification electrodes Y1 - 2002 SN - 0-7484-0791-X ER - TY - BOOK A1 - Wollenberger, Ursula A1 - Renneberg, Reinhard A1 - Bier, Frank Fabian A1 - Scheller, Frieder W. T1 - Analytische Biochemie : eine praktische Einführung in das Messen mit Biomolekülen Y1 - 2003 SN - 3-527-30166-6 PB - John Wiley & Sons CY - Hoboken ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Bier, Frank Fabian T1 - Biosensoren Y1 - 2003 ER - TY - JOUR A1 - Ehrentreich-Förster, Eva A1 - Scheller, Frieder W. A1 - Bier, Frank Fabian T1 - Detection of progesterone in whole blood samples N2 - The progesterone concentration in blood samples can be utilised as a marker for the diagnosis of early pregnancy, endocrinopathy and virilism. Here, we describe a method for progesterone detection and measurement in whole blood samples by a surface sensitive biosensor used in conjunction with an integrated optical grating coupler. This device determines refractive index changes near the biosensor's surface. Hence, biological species bound to a surface layer can be measured in real-time without any label. For the measurements, we have modified the indirect competitive immonoassay principle. The concentration of the progesterone antibody was kept at 1 µg/ml. Progesterone concentration was determined in buffer solution and whole blood in a range between 0.005 and 10 ng/ml. The detection limit was determined to be 3 pM. The relative standard deviation was calculated to be 3.5%. Y1 - 2003 ER - TY - JOUR A1 - Scheller, Frieder W. A1 - Bier, Frank Fabian T1 - Analytical Biochemistry (Editorial) Y1 - 2004 ER - TY - JOUR A1 - Steffen, Jenny A1 - von Nickisch-Rosenegk, Markus A1 - Bier, Frank Fabian T1 - In vitro transcription of a whole gene on a surface-coupled template N2 - An artificial gene was constructed combining the T7 promoter and terminator with the EGFP-gene from the plasmid pEGFP. The functionality of the construct was shown by in vitro translation. The gene-construct was immobilised on a planar glass surface. The transcription was performed on the immobilised gene and mRNA was determined by RT-PCR. Multiple use of the immobilised gene was demonstrated Y1 - 2005 SN - 1473-0189 ER - TY - JOUR A1 - Heise, Christian A1 - Bier, Frank Fabian T1 - Immobilization of DNA on microarrays N2 - Microarrays are new analytical devices that allow the parallel and simultaneous detection of thousands of target compounds. Microarrays, also called DNA chips, are widely used in gene expression, the genotyping of individuals, point mutations, detection of single nucleotide polymorphisms, and short tandem repeats. Microarrays have highly specific base-pair interactions with labeled complementary strands, which makes this technology to a powerful analytical device for monitoring whole genomes. In this article, we provide a survey of the common microarray manufacturing methods, from the selection of support material to surface structuring, immobilization and hybridization, and finally the detection with labeled complementary strands. Special attention is given to the immobilization of single strands, since fast chemical reactions, the creation of homogeneous surface functionalities as well as an oriented coupling are crucial pre-conditions for a good spot morphology and microarrays of high quality Y1 - 2005 ER -