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Karlsruhe Institute of Technology

13 ARTICLES PUBLISHED IN JoVE

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Biology

Chip-based Three-dimensional Cell Culture in Perfused Micro-bioreactors
Eric Gottwald 1, Brigitte Lahni 1, David Thiele 1, Stefan Giselbrecht 1, Alexander Welle 1, Karl-Friedrich Weibezahn 1
1Institute for Biological Interfaces, Forschungszentrum Karlsruhe

We describe a chip-based platform for the three-dimensional cultivation of cells in micro-bioreactors. One chip can house up to 10 Mio. cells that can be cultivated under precisely defined conditions with regard to fluid flow, oxygen tension etc. in a sterile, closed circulation loop.

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Biology

Microfabrication of Chip-sized Scaffolds for Three-dimensional Cell cultivation
Stefan Giselbrecht 1, Eric Gottwald 1, Roman Truckenmueller 2, Christina Trautmann 3, Alexander Welle 1, Andreas Guber 4, Volker Saile 4, Thomas Gietzelt 5, Karl-Friedrich Weibezahn 1
1Institute for Biological Interfaces, Karlsruhe Research Centre, 2Institute for BioMedical Technology, University of Twente, 3Department of Materials Research, Institute for Heavy Ion Research, 4Institute of Microstructure Technology, Karlsruhe Research Centre, 5Institute for Micro Process Engineering, Karlsruhe Research Centre

We present two processes for the microfabrication of porous polymer chips for three-dimensional cell cultivation. The first one is hot embossing combined with a solvent vapour welding process. The second one uses a recently developed microthermoforming process combined with ion track technology leading to a significant simplification of manufacture.

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Biology

Interview: Bioreactors and Surfaced-Modified 3D-Scaffolds for Stem Cell Research
Karl-Friedrich Weibezahn 1
1Institute for Biological Interfaces, Karlsruhe Institute of Technology

In the past many in vitro culture systems -- mainly monolayer cultures -- often suffered from the disadvantage that differentiated primary cells had a relatively short life-span and de-differentiated during culture. As a consequence, most of their organ-specific functions were lost rapidly. Thus, in order to reproduce better conditions for these cells in vitro, modifications and adaptations have been made to conventional monolayer cultures.

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Biology

Intracellular Refolding Assay
Tamara Vanessa Walther 1, Danilo Maddalo 1
1Institute of Toxicology and Genetics, Karlsruhe Institute of Technology

In this protocol a method to measure intracellular protein refolding after heat shock is described. This method can be used to study foldases like molecular chaperones and their co-factors or compounds able to influence their activity. Firefly luciferase activity is used as reporter to measure chaperone refolding activity.

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Bioengineering

Nanotopology of Cell Adhesion upon Variable-Angle Total Internal Reflection Fluorescence Microscopy (VA-TIRFM)
Michael Wagner 1, Petra Weber 1, Harald Baumann 1, Herbert Schneckenburger 1
1Hochschule Aalen, Institut für Angewandte Forschung

Topology of cell adhesion on a substrate is measured with nanometre precision by variable-angle total internal reflection fluorescence microscopy (VA-TIRFM).

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JoVE Core

Stab Wound Injury of the Zebrafish Adult Telencephalon: A Method to Investigate Vertebrate Brain Neurogenesis and Regeneration
Rebecca Schmidt 1, Tanja Beil 1, Uwe Strähle 1, Sepand Rastegar 1
1Institute of Toxicology and Genetics, Karlsruhe Institute of Technology

To shed light on the cellular and molecular mechanisms of zebrafish adult neurogenesis and regeneration, we developed a protocol for invasive surgery causing mechanical injuries in the zebrafish adult telencephalon and subsequent monitoring of changes in the stabbed hemisphere by immunohistochemistry or in situ hybridization.

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Chemistry

Layer-by-layer Synthesis and Transfer of Freestanding Conjugated Microporous Polymer Nanomembranes
Peter Lindemann 1, Yannick Träutlein 1, Christof Wöll 1, Manuel Tsotsalas 1
1Institute of Functional Interfaces, Karlsruhe Institute of Technology

In this paper we describe the interfacial synthesis of conjugated microporous polymers (CMP) on sacrificial substrates, and the dissolution of the substrate for the preparation of freestanding CMP nanomembranes. In addition, we will describe how the fragile nanomembranes can be transferred to other substrates.

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JoVE Journal

Synthesis of Wavelength-shifting DNA Hybridization Probes by Using Photostable Cyanine Dyes
Stefanie Arndt *1, Heidi-Kristin Walter *1, Hans-Achim Wagenknecht 1
1Institute of Organic Chemistry, Karlsruhe Institute of Technology

Photostable cyanine dyes are attached to oligonucleotides to monitor hybridization by energy transfer.

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Genetics

Genetic Manipulation of the Plant Pathogen Ustilago maydis to Study Fungal Biology and Plant Microbe Interactions
Kristin Bösch *1,2, Lamprinos Frantzeskakis *1, Miroslav Vraneš 3, Jörg Kämper 3, Kerstin Schipper 1,2, Vera Göhre 1,2,4
1Institute for Microbiology, Heinrich-Heine University Düsseldorf, 2Bioeconomy Science Center (BioSC), 3Department of Genetics, Institute of Applied Biosciences, Karlsruhe Institute of Technology, 4Cluster of Excellence in Plant Sciences (CEPLAS), Heinrich-Heine University Düsseldorf

We describe a robust gene replacement strategy to genetically manipulate the smut fungus Ustilago maydis. This protocol explains how to generate deletion mutants to investigate infection phenotypes. It can be extended to modify genes in any desired way, e.g., by adding a sequence encoding a fluorescent protein tag.

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Engineering

Challenges in Rheological Characterization of Highly Concentrated Suspensions — A Case Study for Screen-printing Silver Pastes
Ceren Yüce 1, Norbert Willenbacher 1
1Applied Mechanics Group, Institute for Mechanical Process Engineering and Mechanics, Department of Chemical Engineering, Karlsruhe Institute of Technology

A protocol for a robust and application relevant rheological characterization of highly concentrated suspensions is presented. Silver pastes used for screen-printing application in solar cell production are employed as model systems.

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JoVE Journal

Synthesis of Bimetallic Pt/Sn-based Nanoparticles in Ionic Liquids
Christine Dietrich 1,2, Georgios Uzunidis 1,2, Yannick Träutlein 1, Silke Behrens 1,2
1Karlsruhe Institute of Technology (KIT), 2Ruprecht-Karls University of Heidelberg

A protocol for the synthesis of bimetallic nanoparticles in ionic liquids and the procedure of their catalytic testing in the selective hydrogenation of unsaturated aldehydes are described.

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Environment

Automated 3D Optical Coherence Tomography to Elucidate Biofilm Morphogenesis Over Large Spatial Scales
Anna Depetris 1, Antoine Wiedmer 2, Michael Wagner 3, Sebastian Schäfer 4, Tom J. Battin 1, Hannes Peter 1
1Stream Biofilm and Ecosystem Research Laboratory, School of Architecture, Civil and Environmental Engineering, Ecole Polytechnique Fédérale de Lausanne, 2Plateforme technique, School of Architecture, Civil and Environmental Engineering, Ecole Polytechnique Fédérale de Lausanne, 3Water Chemistry and Water Technology, Engler-Bunte-Institut, Karlsruhe Institute of Technology, 4Thorlabs GmbH

Microbial biofilms form complex architectures at interphases and develop into highly scale-dependent spatial patterns. Here, we introduce an experimental system (hard- and software) for the automated acquisition of 3D optical coherence tomography (OCT) datasets. This toolset allows the non-invasive and multi-scale characterization of biofilm morphogenesis in space and time.

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Chemistry

In Vitro and In Vivo Evaluation of Photocontrolled Biologically Active Compounds - Potential Drug Candidates for Cancer Photopharmacology
Kateryna Horbatok *1,2, Tetyana Makhnii *2, Viktoriia Kosach 2, Volodymyr Danko 1, Andrey Kovalenko 1, Stanislav Fatiushchenkov 1, Petro Borysko 2, Iryna Pishel 2, Oleg Babii 3,4, Anne S. Ulrich 3, Tim Schober 4, Sergii Afonin 3, Igor V. Komarov 1,2,4
1Taras Shevchenko National University of Kyiv, 2Bienta/Enamine, 3Karlsruhe Institute of Technology, 4Lumobiotics

This protocol presents a set of experiments adopted for the evaluation of photoswitchable anticancer peptides, that can be used in the preclinical screening of such compounds. This includes cytotoxicity assessment in 2D and 3D cell cultures, the evaluation of ex vivo (model tissue) photoisomerization efficiency, and in vivo efficacy.

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