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Krejci I.,University of Vienna | Piana C.,University of Vienna | Howitz S.,GeSiM MbH | Wegener T.,GeSiM MbH | And 12 more authors.
Acta Biomaterialia | Year: 2012

There is increasing demand for automated cell reprogramming in the fields of cell biology, biotechnology and the biomedical sciences. Microfluidic-based platforms that provide unattended manipulation of adherent cells promise to be an appropriate basis for cell manipulation. In this study we developed a magnetically driven cell carrier to serve as a vehicle within an in vitro environment. To elucidate the impact of the carrier on cells, biocompatibility was estimated using the human adenocarcinoma cell line Caco-2. Besides evaluation of the quality of the magnetic carriers by field emission scanning electron microscopy, the rate of adherence, proliferation and differentiation of Caco-2 cells grown on the carriers was quantified. Moreover, the morphology of the cells was monitored by immunofluorescent staining. Early generations of the cell carrier suffered from release of cytotoxic nickel from the magnetic cushion. Biocompatibility was achieved by complete encapsulation of the nickel bulk within galvanic gold. The insulation process had to be developed stepwise and was controlled by parallel monitoring of the cell viability. The final carrier generation proved to be a proper support for cell manipulation, allowing proliferation of Caco-2 cells equal to that on glass or polystyrene as a reference for up to 10 days. Functional differentiation was enhanced by more than 30% compared with the reference. A flat, ferromagnetic and fully biocompatible carrier for cell manipulation was developed for application in microfluidic systems. Beyond that, this study offers advice for the development of magnetic cell carriers and the estimation of their biocompatibility. © 2011 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.


Kohler S.,University of Leipzig | Weilbeer C.,University of Leipzig | Howitz S.,GeSiM MbH | Becker H.,Microfluidic ChipShop GmbH | And 2 more authors.
Lab on a Chip - Miniaturisation for Chemistry and Biology | Year: 2011

In this work, a microfluidic free-flow electrophoresis device with a novel approach for preventing gas bubbles from entering the separation area is presented. This is achieved by integrating partitioning bars to reduce the channel depth between electrode channels and separation chamber in order to obtain electrical contact and simultaneously prevent bubbles from entering the separation area. The three-layer sandwich chip features a reusable carrier plate with integrated ports for fluidic connection combined with a softlithographically cast microfluidic PDMS layer and a sealing glass slide. This design allows for a straightforward and rapid chip prototyping process. The performance of the device is demonstrated by free-flow zone electrophoretic separations of fluorescent dye mixtures as well as by the separation of labeled amines and amino acids with separation voltages up to 297 V. © 2011 The Royal Society of Chemistry.


Kohler S.,University of Leipzig | Becker H.,Microfluidic ChipShop GmbH | Beushausen V.,Laser Laboratorium Gottingen e.V. | Huttner W.,Laser Laboratorium Gottingen e.V. | And 4 more authors.
Proceedings of SPIE - The International Society for Optical Engineering | Year: 2011

In this work we present the first approach towards low-cost free-flow electrophoresis (FFE) devices utilizing injection molding as a microfabrication process which has the potential to manufacture FFE chips at a cost which make their use commercially viable. This is achieved by realizing a new straightforward micro free-flow electrophoresis (μFFE) design ensuring both, bubble free electrophoretic separation and effective electrical connection by implementing miniaturized partitioning bars. This creates a defined open gap of 20 μm in height and 500 μm in width between separation zone and electrode channels. The thermoplastic μFFE chips are ready to use, there is no need for a subsequent labor-intensive implementation of membranes or salt bridges to separate the electrode channels from the separation zone. © 2011 Copyright Society of Photo-Optical Instrumentation Engineers (SPIE).


Uhlig K.,Fraunhofer Institute for Cell Therapy and Immunology | Wegener T.,GeSiM mbH | He J.,GeSiM mbH | Zeiser M.,Bielefeld University | And 9 more authors.
Biomacromolecules | Year: 2016

Cultivation of adherently growing cells in artificial environments is of utmost importance in medicine and biotechnology to accomplish in vitro drug screening or to investigate disease mechanisms. Precise cell manipulation, like localized control over adhesion, is required to expand cells, to establish cell models for novel therapies and to perform noninvasive cell experiments. To this end, we developed a method of gentle, local lift-off of mammalian cells using polymer surfaces, which are reversibly and repeatedly switchable between a cell-attractive and a cell-repellent state. This property was introduced through micropatterned thermoresponsive polymer coatings formed from colloidal microgels. Patterning was obtained through automated nanodispensing or microcontact printing, making use of unspecific electrostatic interactions between microgels and substrates. This process is much more robust against ambient conditions than covalent coupling, thus lending itself to up-scaling. As an example, wound healing assays were accomplished at 37 °C with highly increased precision in microfluidic environments. (Figure Presented). © 2016 American Chemical Society.


PubMed | Fraunhofer Institute for Cell Therapy and Immunology, Bielefeld University, University of Bayreuth, Leibniz Institute of Polymer Research and GeSiM mbH
Type: Journal Article | Journal: Biomacromolecules | Year: 2016

Cultivation of adherently growing cells in artificial environments is of utmost importance in medicine and biotechnology to accomplish in vitro drug screening or to investigate disease mechanisms. Precise cell manipulation, like localized control over adhesion, is required to expand cells, to establish cell models for novel therapies and to perform noninvasive cell experiments. To this end, we developed a method of gentle, local lift-off of mammalian cells using polymer surfaces, which are reversibly and repeatedly switchable between a cell-attractive and a cell-repellent state. This property was introduced through micropatterned thermoresponsive polymer coatings formed from colloidal microgels. Patterning was obtained through automated nanodispensing or microcontact printing, making use of unspecific electrostatic interactions between microgels and substrates. This process is much more robust against ambient conditions than covalent coupling, thus lending itself to up-scaling. As an example, wound healing assays were accomplished at 37 C with highly increased precision in microfluidic environments.


Markovitz-Bishitz Y.,Bar - Ilan University | Tauber Y.,Bar - Ilan University | Afrimzon E.,Bar - Ilan University | Zurgil N.,Bar - Ilan University | And 5 more authors.
Biomaterials | Year: 2010

Multicellular spheroid models have been recognized as superior to monolayer cell cultures in antitumor drug screening, but their commercial adaptation in the pharmaceutical industry has been delayed, primarily due to technological limitations. The current study presents a new spheroid culture platform that addresses these technical restrictions. The new culturing device is based on a multiwell plate equipped with a glass bottom patterned with an array of UV adhesive microchambers. Each microchamber is designed to accommodate a single spheroid. The system facilitates the simultaneous creation and culturing of a large number of spheroids, as well as screening their response to antitumor drugs. The volume of the spheroids is easily controlled by seeding density. The location of each spheroid is preserved in the same microchamber throughout its growth, treatment with soluble agents, and imaging. The growth ratio parameter, a non-intrusive size analysis of the same spheroid before and after exposure to drugs, was found to be a sensitive indicator for the reaction of MCF7 breast cancer spheroids to cytotoxic drugs. This feature helps reveal the heterogeneity within the spheroid population during the formation process and their drug response, and provides an opportunity to detect specific, highly active or drug-resistant spheroid sub-groups. The advantages of this spheroid-based system make it an efficient drug-screening tool that may be valuable to related fields of research and clinical applications. © 2010 Elsevier Ltd.


Deutsch M.,Bar - Ilan University | Afrimzon E.,Bar - Ilan University | Namer Y.,Bar - Ilan University | Shafran Y.,Bar - Ilan University | And 10 more authors.
BMC Cell Biology | Year: 2010

Background: Cryopreservation is the only widely applicable method of storing vital cells for nearly unlimited periods of time. Successful cryopreservation is essential for reproductive medicine, stem cell research, cord blood storage and related biomedical areas. The methods currently used to retrieve a specific cell or a group of individual cells with specific biological properties after cryopreservation are quite complicated and inefficient.Results: The present study suggests a new approach in cryopreservation, utilizing the Individual Cell-based Cryo-Chip (i3C). The i3C is made of materials having appropriate durability for cryopreservation conditions. The core of this approach is an array of picowells, each picowell designed to maintain an individual cell during the severe conditions of the freezing - thawing cycle and accompanying treatments. More than 97% of cells were found to retain their position in the picowells throughout the entire freezing - thawing cycle and medium exchange. Thus the comparison between pre-freezing and post-thawing data can be achieved at an individual cell resolution. The intactness of cells undergoing slow freezing and thawing, while residing in the i3C, was found to be similar to that obtained with micro-vials. However, in a fast freezing protocol, the i3C was found to be far superior.Conclusions: The results of the present study offer new opportunities for cryopreservation. Using the present methodology, the cryopreservation of individual identifiable cells, and their observation and retrieval, at an individual cell resolution become possible for the first time. This approach facilitates the correlation between cell characteristics before and after the freezing - thawing cycle. Thus, it is expected to significantly enhance current cryopreservation procedures for successful regenerative and reproductive medicine. © 2010 Deutsch et al; licensee BioMed Central Ltd.


Afrimzon E.,Bar - Ilan University | Zurgil N.,Bar - Ilan University | Shafran Y.,Bar - Ilan University | Ehrhart F.,Fraunhofer Institute for Biomedical Engineering | And 11 more authors.
BMC Cell Biology | Year: 2010

Background: The cryopreservation and thawing processes are known to induce many deleterious effects in cells and might be detrimental to several cell types. There is an inherent variability in cellular responses among cell types and within individual cells of a given population with regard to their ability to endure the freezing and thawing process. The aim of this study was to evaluate the fate of cryopreserved cells within an optical cryo apparatus, the individual-cell-based cryo-chip (i3C), by monitoring several basic cellular functional activities at the resolution of individual cells.Results: In the present study, U937 cells underwent the freezing and thawing cycle in the i3C device. Then a panel of vital tests was performed, including the number of dead cells (PI staining), apoptotic rate (Annexin V staining), mitochondrial membrane potential (TMRM staining), cytoplasm membrane integrity and intracellular metabolism (FDA staining), as well as post-thawing cell proliferation assays. Cells that underwent the freezing - thawing cycle in i3C devices exhibited the same functional activity as control cells. Moreover, the combination of the multi-parametric analysis at a single cell resolution and the optical and biological features of the device enable an accurate determination of the functional status of individual cells and subsequent retrieval and utilization of the most valuable cells.Conclusions: The means and methodologies described here enable the freezing and thawing of spatially identifiable cells, as well as the efficient detection of viable, specific, highly biologically active cells for future applications. © 2010 Afrimzon et al; licensee BioMed Central Ltd.


Sonntag F.,Fraunhofer Institute for Material and Beam Technology | Schilling N.,Fraunhofer Institute for Material and Beam Technology | Mader K.,Fraunhofer Institute for Material and Beam Technology | Gruchow M.,Fraunhofer Institute for Material and Beam Technology | And 8 more authors.
Journal of Biotechnology | Year: 2010

Dynamic miniaturized human multi-micro-organ bioreactor systems are envisaged as a possible solution for the embarrassing gap of predictive substance testing prior to human exposure. A rational approach was applied to simulate and design dynamic long-term cultures of the smallest possible functional human organ units, human " micro-organoids" , on a chip the shape of a microscope slide. Each chip contains six identical dynamic micro-bioreactors with three different micro-organoid culture segments each, a feed supply and waste reservoirs. A liver, a brain cortex and a bone marrow micro-organoid segment were designed into each bioreactor. This design was translated into a multi-layer chip prototype and a routine manufacturing procedure was established. The first series of microscopable, chemically resistant and sterilizable chip prototypes was tested for matrix compatibility and primary cell culture suitability. Sterility and long-term human cell survival could be shown. Optimizing the applied design approach and prototyping tools resulted in a time period of only 3 months for a single design and prototyping cycle. This rapid prototyping scheme now allows for fast adjustment or redesign of inaccurate architectures. The designed chip platform is thus ready to be evaluated for the establishment and maintenance of the human liver, brain cortex and bone marrow micro-organoids in a systemic microenvironment. © 2010 Elsevier B.V.

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