Robert Koch-Institute View Institution's Website 4 articles published in JoVE Biology Culture of Piglet Intestinal 3D Organoids from Cryopreserved Epithelial Crypts and Establishment of Cell Monolayers Eloïse Mussard1,2, Corinne Lencina1, Gaëlle Boudry3, Caroline S. Achard2, Christian Klotz4, Sylvie Combes1, Martin Beaumont1 1GenPhySE, Université de Toulouse, INRAE, ENVT, 2Lallemand Animal Nutrition, 3Institut NuMeCan, INRAE, INSERM, Université de Rennes, 4FG 16: Mycotic and Parasitic Agents and Mycobacteria, Robert Koch-Institute Here, we describe a protocol to culture pig intestinal 3D organoids from cryopreserved epithelial crypts. We also describe a method to establish cell monolayers derived from 3D organoids, allowing access to the apical side of epithelial cells. Immunology and Infection Optimized Interferon-gamma ELISpot Assay to Measure T Cell Responses in the Guinea Pig Model after Vaccination Katherine Schultheis1, Hubert Schaefer2, Holly M. Pugh1, Bryan S. Yung1, Janet Oh1, Jacklyn Nguyen1, Laurent Humeau1, Kate E. Broderick1, Trevor R.F. Smith1 1Inovio Pharmaceuticals, Inc, 2Mycotic and Parasitic Agents and Mycobacteria, Robert Koch Institute The development of the interferon-gamma ELISpot assay for guinea pig PBMCs allows the characterization of T-cell responses in this highly relevant model for studying infectious diseases. We have applied the assay to measure T cell responses associated with intradermal delivery of DNA vaccines. Biology Investigating the Detrimental Effects of Low Pressure Plasma Sterilization on the Survival of Bacillus subtilis Spores Using Live Cell Microscopy Felix M. Fuchs1, Marina Raguse1,2,3, Marcel Fiebrandt2, Kazimierz Madela4, Peter Awakowicz2, Michael Laue4, Katharina Stapelmann3, Ralf Moeller1 1Department of Radiation Biology, Institute of Aerospace Medicine, Space Microbiology Research Group, German Aerospace Center (DLR e.V.), 2Institute of Electrical Engineering and Plasma Technology, Faculty of Electrical Engineering and Information Technology, Ruhr-University Bochum, 3Institute of Electrical Engineering and Plasma Technology, Faculty of Electrical Engineering and Information Technology, Biomedical Applications of Plasma Technology, Ruhr-University Bochum, 4Advanced Light and Electron Microscopy (ZBS 4), Robert Koch Institute This protocol illustrates the important consecutive steps required to assess the relevance of monitoring vitality parameter and DNA repair processes in reviving Bacillus subtilis spores after treatment with low pressure plasma by tracking fluorescence-labelled DNA repair proteins via time-resolved confocal microscopy and scanning electron microscopy. Immunology and Infection IgY Technology: Extraction of Chicken Antibodies from Egg Yolk by Polyethylene Glycol (PEG) Precipitation Diana Pauly1, Pablo A. Chacana2, Esteban G. Calzado3, Björn Brembs4, Rüdiger Schade5 1Center for Biological Security, Robert Koch-Institute, 2CICVyA - INTA Castelar, Instituto de Virología, 3Center of Molecular Immunology, Ciudad de la Habana, Cuba, 4Department of Biology, Chemistry, Pharmacy, Institute of Biology-Neurobiology, Free University of Berlin, 5Institut of Pharmacology, Charité-University Medicine of Berlin This protocol describes in particular the extraction of total IgY from egg yolk by means of polyethylene glycol precipitation and gives general information about IgY technology.