Chimera Biotec GmbH

Dortmund, Germany

Chimera Biotec GmbH

Dortmund, Germany
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Schroder H.,Chimera Biotec GmbH | Grosche M.,Karlsruhe Institute of Technology | Adler M.,Chimera Biotec GmbH | Spengler M.,Chimera Biotec GmbH | Niemeyer C.M.,Karlsruhe Institute of Technology
Biochemical and Biophysical Research Communications | Year: 2017

Immuno-PCR (IPCR) combines the versatile ELISA antigen detection with ultrasensitive PCR signal amplification, thereby enabling the highly sensitive detection of a broad range of targets with a typically very large dynamic detection range. The quantification of the antigen is usually achieved by real-time PCR, which provides a correlation between the target concentration and amplified DNA marker. We here report on the implementation of digital droplet PCR as a means for direct quantification of DNA copies to enable the highly sensitive detection of protein biomarkers. To this end, two alternative approaches, based on either magnetic microbead-based IPCR or a microplate-release IPCR were tested. The latter format worked well and revealed an extraordinary high robustness and sensitivity. While rtIPCR already fulfills typical immunoassay acceptance criteria, ddIPCR enables improved accuracy and precision of the assay because signal response and analyte concentrations are directly correlated. The utility of the novel ddIPCR technology is demonstrated at the example of two cytokines, interleukin 2 and interleukin 6 (IL2, IL6, respectively), with an overall average CV% of 5.0 (IL2) and 7.4 (IL6). © 2017 Elsevier Inc.


Diel P.,German Sport University Cologne | Schiffer T.,German Sport University Cologne | Geisler S.,German Sport University Cologne | Hertrampf T.,German Sport University Cologne | And 4 more authors.
Molecular and Cellular Endocrinology | Year: 2010

Myostatin propeptide (MYOPRO) and follistatin (FOLLI) are potent myostatin inhibitors. In this study we analysed effects of training and androgens on MYOPRO and FOLLI concentrations in blood and skeletal muscle using Immuno PCR. Young healthy males performed either a 3-month endurance training or a strength training. Blood and biopsy samples were analysed. Training did not significantly affect MYOPRO and FOLLI concentrations in serum and muscle. To investigate whether total skeletal muscle mass may affect circulating MYOPRO and FOLLI levels, blood samples of tetraplegic patients, untrained volunteers and bodybuilders were analysed. MYOPRO was significantly increased exclusively in the bodybuilder group. In orchiectomised rats MYOPRO increased in blood and muscle after treatment with testosterone. In summary our data demonstrate that moderate training does not affect the concentrations of MYOPRO to FOLLI. In contrast androgen treatment results in a significant increase of MYOPRO in skeletal muscle and serum. © 2010 Elsevier Ireland Ltd.


Leroux-Roels G.,Ghent University | Maes C.,Ghent University | Clement F.,Ghent University | van Engelenburg F.,Kinesis Pharma B.V. | And 9 more authors.
PLoS ONE | Year: 2013

Mucosal antibodies harboring various antiviral activities may best protect mucosal surfaces against early HIV-1 entry at mucosal sites and they should be ideally induced by prophylactic HIV-1 vaccines for optimal prevention of sexually transmitted HIV-1. A phase I, double-blind, randomized, placebo-controlled trial was conducted in twenty-four healthy HIV-uninfected young women. The study objectives were to assess the safety, tolerability and immunogenicity of virosomes harboring surface HIV-1 gp41-derived P1 lipidated peptides (MYM-V101). Participants received placebo or MYM-V101 vaccine at 10 μg/dose or 50 μg/dose intramuscularly at week 0 and 8, and intranasally at week 16 and 24. MYM-V101 was safe and well-tolerated at both doses administered by the intramuscular and intranasal routes, with the majority of subjects remaining free of local and general symptoms. P1-specific serum IgGs and IgAs were induced in all high dose recipients after the first injection. After the last vaccination, vaginal and rectal P1-specific IgGs could be detected in all high dose recipients. Approximately 63% and 43% of the low and high dose recipients were respectively tested positive for vaginal P1-IgAs, while 29% of the subjects from the high dose group tested positive for rectal IgAs. Serum samples had total specific IgG and IgA antibody concentrations ≥0.4 μg/mL, while mucosal samples were usually below 0.01 μg/mL. Vaginal secretions from MYM-V101 vaccinated subjects were inhibiting HIV-1 transcytosis but had no detectable neutralizing activity. P1-specific Th1 responses could not be detected on PBMC. This study demonstrates the excellent safety and tolerability of MYM-V101, eliciting systemic and mucosal antibodies in the majority of subjects. Vaccine-induced mucosal anti-gp41 antibodies toward conserved gp41 motifs were harboring HIV-1 transcytosis inhibition activity and may contribute to reduce sexually-transmitted HIV-1. Trial Registration: ClinicalTrials.gov NCT01084343 http://www.clinicaltrials.gov/ct2/show/NCT01084343?term = NCT01084343&rank = 1. © 2013 Leroux-Roels et al.


Arrabito G.,TU Dortmund | Arrabito G.,Karlsruhe Institute of Technology | Arrabito G.,University of Catania | Reisewitz S.,TU Dortmund | And 9 more authors.
Small | Year: 2013

A general methodology for patterning of multiple protein ligands with lateral dimensions below those of single cells is described. It employs dip pen nanolithography (DPN) patterning of DNA oligonucleotides which are then used as capture strands for DNA-directed immobilization (DDI) of oligonucleotide-tagged proteins. This study reports the development and optimization of PEG-based liquid ink, used as carrier for the immobilization of alkylamino-labeled DNA oligomers on chemically activated glass surfaces. The resulting DNA arrays have typical spot sizes of 4-5 μm with a pitch of 12 μm micrometer. It is demonstrated that the arrays can be further functionalized with covalent DNA-streptavidin (DNA-STV) conjugates bearing ligands recognized by cells. To this end, biotinylated epidermal growth factor (EGF) is coupled to the DNA-STV conjugates, the resulting constructs are hybridized with the DNA arrays and the resulting surfaces used for the culturing of MCF-7 (human breast adenocarcinoma) cells. Owing to the lateral diffusion of transmembrane proteins in the cell's plasma membrane, specific recruitment and concentration of EGF receptor can be induced specifically at the sites where the ligands are bound on the solid substrate. This is a clear demonstration that this method is suitable for precise functional manipulations of subcellular areas within living cells. Direct writing of DNA oligonucleotides by dip pen nanolithography (DPN) and subsequent functionalization of the written patterns with DNA-tagged proteins allows one to produce biochips (red spots) for the precise functional manipulation of subcellular areas within cells, as demonstrated by the recruitment and concentration of transmembrane receptors (green spots) in the plasma membrane of a living cell (light green). Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.


Spengler M.,Chimera Biotec GmbH | Adler M.,Chimera Biotec GmbH | Niemeyer C.M.,Karlsruhe Institute of Technology
Analyst | Year: 2015

Recombinant DNA technology and corresponding innovations in molecular biology, chemistry and medicine have led to novel therapeutic biomacromolecules as lead candidates in the pharmaceutical drug development pipelines. While monoclonal antibodies and other proteins provide therapeutic potential beyond the possibilities of small molecule drugs, the concomitant demand for supportive bioanalytical sample testing creates multiple novel challenges. For example, intact macromolecules can usually not be quantified by mass-spectrometry without enzymatic digestion and isotopically labeled internal standards are costly and/or difficult to prepare. Classical ELISA-type immunoassays, on the other hand, often lack the sensitivity required to obtain pharmacokinetics of low dosed drugs or pharmacodynamics of suitable biomarkers. Here we summarize emerging state-of-the-art ligand-binding assay technologies for pharmaceutical sample testing, which reveal enhanced analytical sensitivity over classical ELISA formats. We focus on immuno-PCR, which combines antibody specificity with the extremely sensitive detection of a tethered DNA marker by quantitative PCR, and alternative nucleic acid-based technologies as well as methods based on electrochemiluminescence or single-molecule counting. Using case studies, we discuss advantages and drawbacks of these methods for preclinical and clinical sample testing. © 2015 The Royal Society of Chemistry.


Kampe T.,TU Dortmund | Konig A.,TU Dortmund | Schroeder H.,TU Dortmund | Schroeder H.,Chimera Biotec GmbH | And 3 more authors.
Analytical Chemistry | Year: 2014

We present a microfluidic device for coupled phase I/phase II metabolic reactions in vitro. The chip consists of microchannels, which are used as packed bed reactor compartments, filled with superparamagnetic microparticles bearing recombinant microsomal phase I cytochrome P450 or phase II conjugating enzymes (UDP-glucuronosyltransferase). Online coupling of the microfluidic device with LC/MS enabled the quantitative assessment of coupled phase I/phase II transformations, as demonstrated for two different substrates, 7-benzyloxy-4-trifluoromethylcoumarin (BFC) and dextromethorphan (DEX). In contrast, conventional sequential one-pot incubations did not generate measurable amounts of phase II metabolites. Because the microfluidic device is readily assembled from standard parts and can be equipped with a variety of recombinant enzymes, it provides a modular platform to emulate and investigate hepatic metabolism processes, with particular potential for targeted small-scale synthesis and identification of metabolites formed by sequential action of specific enzymes. © 2014 American Chemical Society.


Vogel K.,TU Dortmund | Glettenberg M.,TU Dortmund | Schroeder H.,TU Dortmund | Schroeder H.,Chimera Biotec GmbH | And 2 more authors.
Small | Year: 2013

A novel bioorthogonal method for the modification of cells with single-stranded DNA oligomers is compared to five alternative methods with respect to labeling efficacy, specificity, and effects on cell viability. The new method is based on oxime ligation of aminooxybiotin to aldehyde groups installed by periodate cleavage of cell-surface glycans, followed by the coupling of preformed DNA-streptavidin conjugates. As compared with two literature-reported methods based on direct coupling of N-hydroxysuccinimidyl (NHS)-DNA or NHS-biotinylation as well as with techniques based on strain-promoted alkyne-azide cycloaddition, this method shows the highest labeling densities and is sufficiently mild to avoid cell damage. Functionality of the DNA tags is demonstrated by DNA-directed immobilization on solid substrates and assembly of small cell aggregates. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.


Grant
Agency: European Commission | Branch: FP7 | Program: CP-FP | Phase: HEALTH.2010.1.2-1 | Award Amount: 2.57M | Year: 2011

The global frequency of neonatal sepsis is 1-21 newborns per 1000 live births with mortality rates as high as 35-70%, amounting to 100,000s of annual cases. The consortium has identified a real and pressing need for increasing diagnostic efficacy in a setting that imposes a dramatic healthcare burden on EU (and global) healthcare management systems. The goal of this project is the realization and clinical validation of a fully integrated and automated platform for the parallel detection of neonatal sepsis markers and a panel of sepsis-causing bacteria from a clinical sample (whole blood, cerebrospinal fluid, fluid from joint aspirations or urine). For the first time, this will allow a fast (90 minutes) automated routine diagnosis of neonatal sepsis, permitting a timely and especially specific treatment. The project has the potential to significantly reduce neonatal mortality and ensure appropriate treatment for one of the most vulnerable patient cohorts in the medical setting: newborns. The interaction between presence of microbial pathogens and the host (patient) response will be investigated in a clinical validation phase with a view to planning patient management more appropriately. In the long term this will also reduce the spread of multiresistant strains emerging from the unspecific use of broad-spectrum antibiotics. ASCMicroPlat will cover the complete value chain from research and development to clinical validation. The system will be based on the centrifugal microfluidic platform of HSG-IMIT and an inventive foil-technology for the disposable test carriers, which is scalable to industrial production. ASCMicroPlat will enable a sepsis diagnosis based on a sample in result out process. Additionally, the know-how generated in this project will provide innovative European SMEs with a platform for the fast implementation of lab-on-a-chip tests. The consortium has a strong industrial component including 2 SMEs.


Grant
Agency: European Commission | Branch: H2020 | Program: RIA | Phase: NMP-08-2014 | Award Amount: 8.44M | Year: 2015

MACIVIVA is a highly interdisciplinary consortium among well established and innovative SMEs with scientific excellence and complementary industrial world-leading experts with unique expertise and know-how in virosome technology, spray and freeze drying, large scale manufacturing and packaging. MACIVIVA will pave the path to other large scale thermostable nanopharmaceuticals products for therapeutic and prophylactic vaccines and other potential applications for direct application by non-invasive routes. Liquid products are inherently prone to physical and/or chemical modifications and degradations. Solid vaccine dosage formats (e.g. powder) may prevent molecular motion and shear-induced degradation, and slow down degradation involving water and oxygen radicals, resulting in improved stability and enhanced shelf-life of vaccines. The cold chain storage is still fundamental for preserving the bioactivity of most liquid and freeze-dried vaccines, and a reconstitution step prior to administration is required for freeze dried vaccines that are usually administered intramuscularly or subcutaneously. These reconstituted freeze dried vaccines harbor important instability and must be used within hours and kept refrigerated. Because most liquid and reconstituted freeze-dried vaccines are susceptible to degradations, it may affect the immunological properties of the immunogens, with unwanted immune responses or insufficient immune protection. For addressing liquid virosome-based vaccine instability and improving their shelf-life outside the cold chain, MACIVIVA will explore new galenic vaccine formulations through careful screening of excipients, stabilization and drying methods for generating new vaccine solid forms that can be easily self-administered. Robust universal manufacturing processes for upscale production of virosome dried powder for the non-invasive intranasal, oral and sublingual routes should be achieved by month 42.


Patent
Chimera Biotec GmbH | Date: 2010-05-26

The present invention relates to novel conjugate complexes for immunoassays as well as kits comprising these conjugate complexes, methods of producing these complexes, and methods of detecting an analyte by use of these complexes. The conjugate complexes of the invention comprise one or more non-nucleic acid receptors capable of specifically binding an analyte, one or more nucleic acid markers comprising a predetermined nucleotide sequence, one or more first linker molecules capable of specifically binding the non-nucleic acid receptor and the nucleic acid marker, and one or more second linker molecules capable of specifically binding the first linker molecules.

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