Fraunhofer Institute for Biomedical Engineering


Fraunhofer Institute for Biomedical Engineering

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News Article | April 29, 2016

Tiny dwarves keep our mattresses clean, repair damage to our teeth, stop eggs sticking to our pans, and extend the shelf life of our food. We are talking about nanomaterials – "nano" comes from the Greek word for "dwarf". These particles are just a few billionths of a meter small, and they are used in a wide range of consumer products. However, up to now the impact of these materials on the environment has been largely unknown, and information is lacking on the concentrations and forms in which they are present there. "It's true that many laboratory studies have examined the effect of nanomaterials on human and animal cells. To date, though, it hasn't been possible to detect very small amounts in environmental samples," says Dr. Yvonne Kohl from the Fraunhofer Institute for Biomedical Engineering IBMT in Sulzbach. That is precisely the objective of the NanoUmwelt project. The interdisciplinary project team is made up of eco- and human toxicologists, physicists, chemists and biologists, and they have just managed to take their first major step forward in achieving their goal: they have developed a method for testing a variety of environmental samples such as river water, animal tissue, or human urine and blood that can detect nanomaterials at a concentration level of nanogram per liter (ppb – parts per billion). That is equivalent to half a sugar cube in the volume of water contained in 1000 competition swimming pools. Using the new method, it is now possible to detect not just large amounts of nanomaterials in clear fluids, as was previously the case, but also very few particles in complex substance mixtures such as human blood or soil samples. The approach is based on field-flow fractionation (FFF), which can be used to separate complex heterogeneous mixtures of fluids and particles into their component parts – while simultaneously sorting the key components by size. This is achieved by the combination of a controlled flow of fluid and a physical separation field, which acts perpendicularly on the flowing suspension. For the detection process to work, environmental samples have to be appropriately processed. The team from Fraunhofer IBMT's Bioprocessing & Bioanalytics Department prepared river water, human urine, and fish tissue to be fit in the FFF device. "We prepare the samples with special enzymes. In this process, we have to make sure that the nanomaterials are not destroyed or changed. This allows us to detect the real amounts and forms of the nanomaterials in the environment," explains Kohl. The scientists have special expertise when it comes to providing, processing and storing human tissue samples. Fraunhofer IBMT has been running the "German Environmental Specimen Bank (ESB) – Human Samples" since January 2012 on behalf of Germany's Environment Agency (UBA). Each year the research institute collects blood and urine samples from 120 volunteers in four cities in Germany. Individual samples are a valuable tool for mapping the trends over time of human exposure to pollutants. "In addition, blood and urine samples have been donated for the NanoUmwelt project and put into cryostorage at Fraunhofer IBMT. We used these samples to develop our new detection method," says Dr. Dominik Lermen, manager of the working group on Biomonitoring & Cryobanks at Fraunhofer IBMT. After approval by the UBA, some of the human samples in the ESB archive may also be examined using the new method. Nanomaterials end up in the environment via different pathways, inter alia the sewage system. Human beings and animals presumably absorb them through biological barriers such as the lung or intestine. The project team is simulating these processes in petri dishes in order to understand how nanomaterials are transported across these barriers. "It's a very complex process involving an extremely wide range of cells and layers of tissue," explains Kohl. The researchers replicate the processes in a way as realistic as possible. They do this by, for instance, measuring the electrical flows within the barriers to determine the functionality of these barriers – or by simulating lung-air interaction using clouds of artificial fog. In the first phase of the NanoUmwelt project, the IBMT team succeeded in developing several cell culture models for the transport of nanomaterials across biological barriers. IBMT worked together with the Fraunhofer Institute for Molecular Biology and Applied Ecology IME, which used pluripotent stem cells to develop a model for investigating cardiotoxicity. Empa, the Swiss partner in the project, delivered a placental barrier model for studying the transport of nanomaterials between mother and child. Next, the partners want to use their method to measure the concentrations of nanoparticles in a wide variety of environmental samples. They will then analyze the results obtained so as to be in a better position to assess the behavior of nanomaterials in the environment and their potential danger for humans, animals, and the environment. "Our next goal is to detect particles in even smaller quantities," says Kohl. To achieve this, the scientists are planning to use special filters to remove distracting elements from the environmental samples, and they are looking forward to develop new processing techniques. Explore further: Nanomaterials in our environment

News Article | December 1, 2015

For the drug tests of the future, the pharma industry needs large quantities of pluripotent stem cells. These stem cells have the potential to transform themselves into any kind of somatic cell, such as the cells of inner organs. Many thousands of stem cell lines from a huge variety of patients are currently being built up in biobanks, where doctors can access perfect models of the genetic illnesses of these patients. Using these stem cells, doctors and pharmaceutical companies can test new drugs better and more quickly than before. Scientists at the Fraunhofer Institute for Biomedical Engineering IBMT in Sulzbach have identified seaweed from Chile as a particularly efficient source of nutrients for the expansion of pluripotent stem cells. Over the past few years, they have developed a controlled and documented production process for alginate, the seaweed's supporting structure. The process encompasses everything from harvesting the seaweed on Chilean beaches and in the seas off Chile, to importing the granulated and dried seaweed, to manufacturing the alginate and using it in cell culture to grow pluripotent stem cells at the institute in Saarland. British pharma companies are currently validating the process in their laboratories. "The first concrete trials with partners from the European Federation of Pharmaceutical Industries and Associations (EFPIA) are planned for next year," says Prof. Heiko Zimmermann, Managing Head of Fraunhofer IBMT. "The goal is to demonstrate that we can use the process to produce stable pluripotent stem cells. At the institute, we've already managed to do just that for many individual stem cell lines." The Fraunhofer scientists at Sulzbach developed the production process and the technology platform jointly with their colleagues in Chile and the United Kingdom. Alginate from two Chilean seaweed types particularly suitable Two seaweed species that grow on the coast of Chile form the source material: Lessonia trabeculata and Lessonia nigrescens. Supporting structures in the cell walls of the seaweed are made of alginate, which is particularly suitable for stem cell cultivation: it consists of a highly aqueous gel that is more viscous than honey. When cross-linked with calcium or barium, it is both stable and flexible – like the jello you find in your dessert bowl – and also permeable for nutrients and important factors. "Cells feel especially at home in elastic 3D environments such as are found inside the body. It's precisely this environment that can be simulated perfectly using alginate," explains Prof. Zimmermann. This is an ideal environment particularly for heart muscle cells, which contract regularly. The scientists flexibly set the elasticity through the mixture of seaweed species and produce the alginate in beads of any size. "After all, different cells need different culture conditions," says Prof. Zimmermann. "We also introduce active ingredients into the alginate and release them in a controlled manner." Examples of such ingredients are substances that transform pluripotent stem cells into certain somatic cells. "In the future," continues Prof. Zimmermann, "the alginate will not only act as a passive substrate, but will also actively influence the growth of the stem cells." The absence of autofluorescence in the elastic biomass is a further advantage and is important for optical analysis techniques. "The stem cells grow better on our alginate – and particularly well in automated bioreactors. They differentiate better into the desired somatic cells than on the plastic substrates generally used today," says Prof. Zimmermann. Harvesting the seaweed is subject to rigorous controls: there are special licenses for Chilean fishers, who harvest only the seaweed that is suitable for manufacturing the alginate, and only as much as permits sustainable resource management on the Chilean coast. In a laboratory operated by IBMT and Fraunhofer Chile at UCN University in Coquimbo, the seaweed is individually peeled, shredded, and completely dried. This is all done within 24 hours to prevent the material from becoming contaminated. The seaweed granulate is then exported to Germany, where IBMT scientists separate out the alginate in the institute's cleanroom. After this process, it is available in liquid form and can be shaped into beads using a strong jet of air. "The beads are rendered more stable in a barium bath, as barium tends to remain in the seaweed mass. The trick is to make the material stable, but not too hard," says Prof. Zimmermann. The researchers place the protein-coated alginate into a bioreactor, which provides the optimum temperature and CO2 environment and continuously stirs the nutrients and cells. Measuring around 200 micrometers, each individual alginate bead performs the role of a Petri dish. The stem cells grow over the alginate in the containers in three to seven days, propagating as they do so. "Because the alginate volumes in the reactors can be increased slightly, we can grow pluripotent stem cells in greater quantities and in smaller spaces," says Prof. Zimmermann.

Teller C.,Hebrew University of Jerusalem | Teller C.,Fraunhofer Institute for Biomedical Engineering | Willner I.,Hebrew University of Jerusalem
Trends in Biotechnology | Year: 2010

The structural and functional information encoded in the base sequence of nucleic acids provides a means to organize hybrid protein-DNA nanostructures with pre-designed, programmed functionality. This review discusses the activation of enzyme cascades in supramolecular DNA-protein hybrid structures, the bioelectrocatalytic activation of redox enzymes on DNA scaffolds, and the programmed positioning of enzymes on 1D, 2D and 3D DNA nanostructures. These systems provide starting points towards the design of interconnected enzyme networks. Substantial progress in the tailoring of functional protein-DNA nanostructures has been accomplished in recent years, and advances in this field warrant a comprehensive discussion. The application of these systems for the control of biocatalytic transformations, for amplified biosensing, and for the synthesis of metallic nanostructures are addressed, and future prospects for these systems are highlighted. © 2010 Elsevier Ltd.

Schmidt S.,Max Planck Institute of Colloids and Interfaces | Schmidt S.,Fraunhofer Institute for Biomedical Engineering | Volodkin D.,Fraunhofer Institute for Biomedical Engineering
Journal of Materials Chemistry B | Year: 2013

The article reviews recent studies on template assisted preparation of microparticles composed of proteins. The main line of development is focused on calcium carbonate (CaCO3) as microscopic sacrificial templates due to their biocompatibility and mild decomposition conditions that are mandatory in order to process fragile biomolecules. Consequently, we discuss several recent methods toward the preparation of protein particles via CaCO3 templating as well as synthesis of the templates. We emphasize how the physical and chemical properties of the template and proteins are exploited in order to obtain pure microparticulate protein therapeutics and multicomponent particle systems. We report on strategies that allow the tuning of material properties like catalytic activity, mechanical properties as well as capacity for a pharmaceutical payload. Finally, we give an overview on the perspectives of CaCO3 templated particles in biomedical applications. © 2013 The Royal Society of Chemistry.

Pechenkin M.A.,Moscow State University | Mohwald H.,Max Planck Institute of Colloids and Interfaces | Volodkin D.V.,Fraunhofer Institute for Biomedical Engineering
Soft Matter | Year: 2012

We have studied the pH- and salt-mediated response of matrix-type polyelectrolyte microcapsules. The capsules were prepared by layer-by-layer (LbL) adsorption on decomposable CaCO3 cores using model polyelectrolytes, namely poly-styrenesulfonic acid (PSS) and poly(allylamine hydrochloride) (PAH). Salt-mediated LbL-made microcapsule fusion has been reported recently with a different polycation (R. Zhang, O. Kreft, A. Skirtach, H. Mohwald and G. Sukhorukov, Soft Matter, 2010, 6, 4742-4747) resulting in merging of the capsule's content and formation of anisotropic "Janus-like" capsules indicating no polymer exchange between the capsules. Here we have studied PAH/PSS capsule behavior as a function of pH and salt concentration. Salt (NaCl) does not induce any changes in the capsules up to saturation concentration (6.1 M). In contrast, several sequential processes have been identified for capsules in [HCl] > 0.1 M: (i) shrinkage due to polymer network annealing, (ii) transformation from matrix-type to shell-type capsules due to oscillating inflating-deflating cycles caused by CO2 formation, and (iii) collapse. The processes depend on acid concentration and the number of layers. If the capsules contact each other, there is an exchange of polymer molecules followed by fusion. The polymer exchange depends on the outermost layer that determines the overall capsule charge. Exchange is enhanced for capsules of the same outermost layer and can be caused by charge redistribution in the capsules at low pH (the negative charge of PSS is reduced (pKa 1) and the positive charge of PAH is in excess). Control over polymer exchange between the capsules is key in order to design capsules as well as to understand and to trigger fusion. We also show that the observed processes are not reversible and can be stopped at any time by replacement of acid with water. Stable gas-filled capsules can be produced by this method upon transformation from matrix to shell-type capsules. This journal is © 2012 The Royal Society of Chemistry.

Volodkin D.,Fraunhofer Institute for Biomedical Engineering | Von Klitzing R.,TU Berlin
Current Opinion in Colloid and Interface Science | Year: 2014

The competition of interactions between charged groups of polyanions and polycations and their interaction with small counterions strongly affect the formation and stability of polyelectrolyte multilayers. This has consequences for the properties of polyelectrolyte multilayers like mechanics, polymer mobility and swelling in water. © 2014 Elsevier Ltd.

Volodkin D.,Fraunhofer Institute for Biomedical Engineering
Advances in Colloid and Interface Science | Year: 2014

Porous CaCO3 vaterite microparticles have been introduced a decade ago as sacrificial cores and becoming nowadays as one of the most popular templates to encapsulate bioactive molecules. This is due to the following beneficial features: i) mild decomposition conditions, ii) highly developed surface area, and iii) controlled size as well as easy and chip preparation. Such properties allow one to template and design particles with well tuned material properties in terms of composition, structure, functionality - the parameters crucially important for bioapplications. This review presents a recent progress in utilizing the CaCO3 cores for the assembly of micrometer-sized beads and capsules with encapsulated both small drugs and large biomacromolecules. Bioapplications of all the particles for drug delivery, biotechnology, and biosensing as well as future perspectives for templating are addressed. © 2014 Published by Elsevier B.V.

Volodkin D.,Fraunhofer Institute for Biomedical Engineering
Colloid and Polymer Science | Year: 2014

Colloidal particles from pure proteins are favorable over composite colloids (usually polymer-based) for applications in drug delivery and biocatalysis. This is due to degradation issue and protein unfolding. Hard templating based on porous CaCO3 cores has been recently adopted for fabrication of pure protein colloids. In comparison to conventional techniques, the templating offers (i) a control over particles size and (ii) mild preparation conditions without any additives, shear forces, and exposure to high temperature or gas-water interface. In this review, the current achievements in CaCO3-based templating of protein colloids are given. The focus is on physicochemical and material properties of the colloids such as stability, mechanical properties, and internal structure. These properties are considered as a function of pH, ionic strength, and protein denaturation degree. Understanding of these basic aspects gives an option to formulate the protein colloids by hard templating achieving desired particle properties that is crucially important for future applications. © 2014 Springer-Verlag.

Brodel A.K.,Fraunhofer Institute for Biomedical Engineering | Sonnabend A.,Fraunhofer Institute for Biomedical Engineering | Kubick S.,Fraunhofer Institute for Biomedical Engineering
Biotechnology and Bioengineering | Year: 2014

Protein expression systems are widely used in biotechnology and medicine for the efficient and economic production of therapeutic proteins. Today, cultivated Chinese hamster ovary (CHO) cells are the market dominating mammalian cell-line for the production of complex therapeutic proteins. Despite this outstanding potential of CHO cells, no high-yield cell-free system based on translationally active lysates from these cells has been reported so far. To date, CHO cell extracts have only been used as a foundational research tool for understanding mRNA translation (Lodish et al., 1974; McDowell et al., 1972). In the present study, we address this fact by establishing a novel cell-free protein expression system based on extracts from cultured CHO cells. Lysate preparation, adaptation of in vitro reaction conditions and the construction of particular expression vectors are considered for high-yield protein production. A specific in vitro expression vector, which includes an internal ribosome entry site (IRES) from the intergenic region (IGR) of the Cricket paralysis virus (CrPV), has been constructed in order to obtain optimal performance. The IGR IRES is supposed to bind directly to the eukaryotic 40S ribosomal subunit thereby bypassing the process of translation initiation, which is often a major bottleneck in cell-free systems. The combination of expression vector and optimized CHO cell extracts enables the production of approximately 50μg/mL active firefly luciferase within 4h. The batch-type cell-free coupled transcription-translation system has the potential to perform post-translational modifications, as shown by the glycosylation of erythropoietin. Accordingly, the system contains translocationally active endogenous microsomes, enabling the co-translational incorporation of membrane proteins into biological membranes. Hence, the presented in vitro translation system is a powerful tool for the fast and convenient optimization of expression constructs, the specific labeling of integral membrane proteins and the cell-free production of posttranslationally modified proteins. © 2013 Wiley Periodicals, Inc.

Scheller F.W.,Fraunhofer Institute for Biomedical Engineering
Advances in biochemical engineering/biotechnology | Year: 2014

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.

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