Utrecht University

17 articles published in JoVE

• Bioengineering

  
  Quantifying Cytoskeleton Dynamics Using Differential Dynamic Microscopy Hannah N. Verwei¹, Gloria Lee², Gregor Leech², Irene Istúriz Petitjean³, Gijsje H. Koenderink³, Rae M. Robertson-Anderson², Ryan James McGorty^{2 1}Cell Biology, Neurobiology and Biophysics, Department of Biology, Faculty of Science, Utrecht University, ²Department of Physics and Biophysics, University of San Diego, ³Department of Bionanoscience, Kavli Institute of Nanoscience Delft, Delft University of Technology Differential dynamic microscopy (DDM) combines features of dynamic light scattering and microscopy. Here, the process of using DDM to characterize reconstituted cytoskeleton networks by quantifying the subdiffusive and caged dynamics of particles in vimentin networks and the ballistic motion of active myosin-driven actin-microtubule composites is presented.

• Biochemistry

  
  Monitoring Protein Aggregation Kinetics In Vivo using Automated Inclusion Counting in Caenorhabditis elegans Jelle Molenkamp*¹, Anna den Outer*¹, Vera van Schijndel*¹, Tessa Sinnige^{1 1}Bijvoet Centre for Biomolecular Research, Utrecht University Here, a method is presented for the analysis of protein aggregation kinetics in the nematode Caenorhabditis elegans. Animals from an age-synchronized population are imaged at different time points, followed by semiautomated inclusion counting in CellProfiler and fitting to a mathematical model in AmyloFit.

• Neuroscience

  
  A High-Throughput Image-Guided Stereotactic Neuronavigation and Focused Ultrasound System for Blood-Brain Barrier Opening in Rodents Rianne Haumann*^1,2, Elvin ’t Hart*², Marc P. P. Derieppe², Helena C. Besse³, Gertjan J. L. Kaspers^1,2, Eelco Hoving², Dannis G. van Vuurden^1,2, Esther Hulleman^1,2, Mario Ries^{3 1}Amsterdam UMC, Vrije Universiteit Amsterdam, Pediatric Oncology, Cancer Center Amsterdam, ²Princess Máxima Center for Pediatric Oncology, ³Imaging Division, Utrecht University The blood-brain barrier (BBB) can be temporarily disrupted with microbubble-mediated focused ultrasound (FUS). Here, we describe a step-by-step protocol for high-throughput BBB opening in vivo using a modular FUS system accessible for non-ultrasound experts.

• Medicine

  
  Characterization of Sickling During Controlled Automated Deoxygenation with Oxygen Gradient Ektacytometry Minke A.E. Rab^1,2, Brigitte A. van Oirschot¹, Jennifer Bos¹, Celeste K. Kanne³, Vivien A. Sheehan³, Eduard J. van Beers², Richard van Wijk^{1 1}Laboratory of Clinical Chemistry and Hematology, University Medical Center Utrecht, Utrecht University, ²Van Creveldkliniek, University Medical Center Utrecht, Utrecht University, ³Department of Pediatrics, Division of Hematology/Oncology, Baylor College of Medicine Here, we present oxygen gradient ektacytometry, a rapid and reproducible method to measure red blood cell deformability in samples from patients with sickle cell disease under controlled deoxygenation and reoxygenation. This technique provides a way to study red blood cell sickling and to monitor sickle cell disease treatment efficacy.

• Behavior

  
  Brain Infarct Segmentation and Registration on MRI or CT for Lesion-symptom Mapping J. Matthijs Biesbroek¹, Hugo J. Kuijf², Nick A. Weaver¹, Lei Zhao³, Marco Duering⁴, Meta VCI Map Consortium, Geert Jan Biessels^{1 1}Department of Neurology and Neurosurgery, UMC Utrecht Brain Center, University Medical Center Utrecht, Utrecht University, ²Image Sciences Institute, University Medical Center Utrecht, ³BrainNow Research Institute, ⁴Institute for Stroke and Dementia Research, University Hospital, LMU Munich Provided here is a practical tutorial for an open-access, standardized image processing pipeline for the purpose of lesion-symptom mapping. A step-by-step walkthrough is provided for each processing step, from manual infarct segmentation on CT/MRI to subsequent registration to standard space, along with practical recommendations and illustrations with exemplary cases.

• Biochemistry

  
  Analysis of Protein Folding, Transport, and Degradation in Living Cells by Radioactive Pulse Chase Nicholas McCaul^1,2, Hui Ying Yeoh¹, Guus van Zadelhoff¹, Naomi Lodder¹, Bertrand Kleizen¹, Ineke Braakman^{1 1}Cellular Protein Chemistry, Utrecht University, ² Here we describe a protocol for a general pulse-chase method that allows the kinetic analysis of folding, transport, and degradation of proteins to be followed in live cells.

• Bioengineering

  
  Fabrication of Decellularized Cartilage-derived Matrix Scaffolds Kim E.M. Benders¹, Margo L. Terpstra¹, Riccardo Levato¹, Jos Malda^{1,2 1}Department of Orthopedics, University Medical Center Utrecht, The Netherlands, ²Department of Veterinary Medicine, Equine Surgery, University of Utrecht, The Netherlands Decellularized cartilage-derived scaffolds can be used as a scaffold to guide cartilage repair and as a means to regenerate osteochondral tissue. This paper describes the decellularization process in detail and provides suggestions to use these scaffolds in in vitro settings.

• Medicine

  
  Transplantation of Adipose Tissue-Derived Stem Cell Sheet to Reduce Leakage After Partial Colectomy in A Rat Model Panithi Sukho^1,2,3, Geesien S.A. Boersema⁴, Nicole Kops⁵, Johan F. Lange⁴, Jolle Kirpensteijn^1,6, Jan Willem Hesselink¹, Yvonne M. Bastiaansen-Jenniskens⁵, Femke Verseijden^{1,5 1}Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, ²Department of Otorhinolaryngology, Erasmus MC University Medical Center, ³Department of Clinical Sciences and Public Health, Faculty of Veterinary Science, Mahidol University, ⁴Department of Surgery, Erasmus MC University Medical Center, ⁵Department of Orthopaedics, Erasmus MC University Medical Center, ⁶ The preparation and transplantation of adipose tissue derived stem cell (ASC) sheets onto insufficiently sutured colorectal anastomoses in a rat model is presented. This novel application shows that ASC sheets can reduce colorectal anastomosis leakage.

• Bioengineering

  
  Microfluidic Bioprinting for Engineering Vascularized Tissues and Organoids Yu Shrike Zhang*¹, Qingmeng Pi*^1,2, Anne Metje van Genderen^{1,3 1}Division of Engineering in Medicine, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, ²Department of Plastic and Reconstructive Surgery, Renji Hospital, Shanghai Jiao Tong University School of Medicine, ³Division of Pharmacology, Utrecht Institute for Pharmaceutical Sciences, Utrecht University We provide a generalized protocol based on a microfluidic bioprinting strategy for engineering a microfibrous vascular bed, where a secondary cell type could be further seeded into the interstitial space of this microfibrous structure to generate vascularized tissues and organoids.

• Biology

  
  Live-cell Imaging of Platelet Degranulation and Secretion Under Flow Arjan D. Barendrecht*¹, Johan J. F. Verhoef*², Silvia Pignatelli¹, Gerard Pasterkamp¹, Harry F. G. Heijnen¹, Coen Maas^{1 1}Department of Clinical Chemistry and Haematology, University Medical Center Utrecht, ²Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University This work describes a fluorescence microscopy-based method for the study of platelet adhesion, spreading, and secretion under flow. This versatile platform enables the investigation of platelet function for mechanistic research on thrombosis and hemostasis.

• Medicine

  
  Isolation and Culture of Primary Endothelial Cells from Canine Arteries and Veins Loes A. Oosterhoff¹, Hedwig S. Kruitwagen¹, Bart Spee¹, Frank G. van Steenbeek^{1 1}Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University Novel isolation methods of primary endothelial cells from blood vessels are needed. This protocol describes a new technique that completely inverts blood vessels of interest, exposing only the endothelial side to enzymatic digestion. The resulting pure endothelial cell culture can be used to study cardiovascular diseases, disease modelling, and angiogenesis.

• Immunology and Infection

  
  A Miniaturized Glycan Microarray Assay for Assessing Avidity and Specificity of Influenza A Virus Hemagglutinins Ryan McBride¹, James C. Paulson¹, Robert P. de Vries^{1,2 1}Department of Cell and Molecular Biology, Chemical Physiology and Microbial Science, The Scripps Research Institute, ²Department of Medicinal Chemistry and Chemical Biology, Utrecht Institute for Pharmaceutical Sciences, Utrecht University Using a printed glycan microarray strategy, a conventional 96-well plate assay was miniaturized for analysis of influenza A virus hemagglutinin avidity and specificity for sialic acid containing receptors.

• Developmental Biology

  
  Imaging Subcellular Structures in the Living Zebrafish Embryo Peter Engerer¹, Gabriela Plucinska^1,2, Rachel Thong^1,3, Laura Trovò¹, Dominik Paquet^4,5,6, Leanne Godinho^{1 1}Institute of Neuronal Cell Biology, Technische Universität München, ²Cell Biology, Department of Biology, Faculty of Science, Utrecht University, ³Faculty of Biology, Ludwig-Maximilians-Universität-München, ⁴Adolf-Butenandt-Institute, Biochemistry, Ludwig-Maximilians-Universität-München, ⁵German Center for Neurodegenerative Diseases, ⁶Laboratory of Brain Development and Repair, The Rockefeller University Imaging the dynamic behavior of organelles and other subcellular structures in vivo can shed light on their function in physiological and disease conditions. Here, we present methods for genetically tagging two organelles, centrosomes and mitochondria, and imaging their dynamics in living zebrafish embryos using wide-field and confocal microscopy.

• Bioengineering

  
  Hybrid µCT-FMT imaging and image analysis Felix Gremse*¹, Dennis Doleschel*¹, Sara Zafarnia¹, Anne Babler², Willi Jahnen-Dechent², Twan Lammers^1,3, Wiltrud Lederle¹, Fabian Kiessling^{1 1}Experimental Molecular Imaging, RWTH Aachen University, ²Institute for Biomedical Engineering - Biointerface Laboratory, RWTH Aachen University, ³Utrecht Institute for Pharmaceutical Sciences, Utrecht University We describe a protocol for hybrid imaging, combining fluorescence-mediated tomography (FMT) with micro computed tomography (µCT). After fusion and reconstruction, we perform interactive organ segmentation to extract quantitative measurements of the fluorescence distribution.

• Behavior

  
  The Modified Hole Board - Measuring Behavior, Cognition and Social Interaction in Mice and Rats Maaike Labots^1,2, Hein A. Van Lith^1,2, Frauke Ohl^1,2, Saskia S. Arndt^{1,2 1}Department of Animals in Science and Society, Utrecht University, ²Brain Center Rudolf Magnus This protocol describes the modified hole board, which is a behavioral test set-up that comprises the characteristics of an open field and a traditional hole board. This set-up enables the differential analysis of unconditioned behavior of small laboratory mammals as well as the analysis of cognitive abilities.

• Neuroscience

  
  Isolation of CA1 Nuclear Enriched Fractions from Hippocampal Slices to Study Activity-dependent Nuclear Import of Synapto-nuclear Messenger Proteins Pingan Yuanxiang¹, Sujoy Bera¹, Anna Karpova¹, Michael R. Kreutz¹, Marina Mikhaylova^{1,2 1}RG Neuroplasticity, Leibniz Institute for Neurobiology, ²Department of Cell Biology, Utrecht University We provide a detailed protocol for induction of long-term potentiation in the CA1 region of the hippocampus and the subsequent isolation of nuclear enriched fractions from the tetanized area of the slice. This approach can be used to determine activity dependent nuclear protein import in cellular models of learning and memory.

• Biology

  
  Sorting of Streptomyces Cell Pellets Using a Complex Object Parametric Analyzer and Sorter Marloes L. C. Petrus¹, G. Jerre van Veluw², Han A. B. Wösten², Dennis Claessen^{1 1}Microbial Biotechnology, Institute Biology Leiden, Leiden University, ²Microbiology, Kluyver Centre for Genomics of Industrial Fermentation, Utrecht University Liquid-grown Streptomyces cultures are characterized by mycelial pellets that are heterogeneous in size. We here describe a method to analyze and sort such pellets in a high-throughput manner. These pellets can be used for further analyses, which will provide leads to understand and control growth heterogeneity.