AP-HP 6 articles published in JoVE Biology Unfractionated Bulk Culture of Mouse Skeletal Muscle to Recapitulate Niche and Stem Cell Quiescence Louai Zaidan*1, Perla Geara*1, Matthew J. Borok*1, Léo Machado1, Despoina Mademtzoglou1, Philippos Mourikis1, Frederic Relaix1,2,3,4 1Univ Paris Est Creteil, INSERM, IMRB, F-94010 Creteil, France, 2Ecole nationale vétérinaire d'Alfort, IMRB, F-94700 Maisons-Alfort, France, 3EFS, IMRB, F-94010 Creteil, France, 4AP-HP, Hopital Mondor, Service d'histologie, F-94010 Creteil, France Skeletal muscle comprises multiple cell types, including resident stem cells, each with a special contribution to muscle homeostasis and regeneration. Here, the 2D culture of muscle stem cells and the muscle cell niche in an ex vivo setting that preserves many of the physiological, in vivo, and environmental characteristics are described. Immunology and Infection Generation, Amplification, and Titration of Recombinant Respiratory Syncytial Viruses Camille Bouillier1, Vincent Rincheval1, Delphine Sitterlin1, Sabine Blouquit-Laye1, Aurore Desquesnes1, Jean-François Eléouët2, Elyanne Gault1,3, Marie-Anne Rameix-Welti1,3 1UMR 1173 Institut national de la santé et de la recherche médicale (INSERM), Université de Versailles St. Quentin, 2UR892 Institut national de la recherche agronomique (INRA), Unité de virologie et immunologie moléculaires, 3Assistance Publique–Hôpitaux de Paris (AP-HP), Hôpital Ambroise Paré, Laboratoire de Microbiologie We describe a method for generating and amplifying genetically modified respiratory syncytial viruses (RSVs) and an optimized plaque assay for RSVs. We illustrate this protocol by creating two recombinant viruses that respectively allow quantification of RSV replication and live analysis of RSV inclusion bodies and inclusion bodies-associated granules dynamics. Cancer Research Immunoglobulin Gene Sequence Analysis In Chronic Lymphocytic Leukemia: From Patient Material To Sequence Interpretation Andreas Agathangelidis*1, Lesley Ann Sutton*2,3, Anastasia Hadzidimitriou1, Cristina Tresoldi4, Anton W. Langerak5, Chrysoula Belessi6, Frederic Davi7, Richard Rosenquist2,3, Kostas Stamatopoulos1,2, Paolo Ghia8 1Institute of Applied Biosciences, Centre for Research and Technology Hellas, 2Department of Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, 3Department of Molecular Medicine and Surgery, Karolinska Institutet, 4Division of Immunology, Transplantation and Infectious, IRCCS San Raffaele Scientific Institute, 5Department of Immunology, Laboratory for Medical Immunology, Erasmus University Medical Center, 6Hematology Department, Nikea General Hospital, 7Assistance publique - Hôpitaux de Paris (AP-HP), Hopital Pitié-Salpêtrière, Department of Hematology, and UPMC University Paris 06, UMRS 1138, 8Division of Experimental Oncology, IRCCS Istituto Scientifico San Raffaele and Università Vita-Salute San Raffaele Herein, we present a protocol that details the technical aspects and essential requirements to ensure robust IG gene sequence analysis in patients with chronic lymphocytic leukemia (CLL), based on the accumulated experience of the European Research initiative on CLL (ERIC). Developmental Biology Engineering Transplantation-suitable Retinal Pigment Epithelium Tissue Derived from Human Embryonic Stem Cells Karim Ben M'Barek1,2,3, Walter Habeler1,2,3, Alexandra Plancheron1,2,3, Mohamed Jarraya4, Olivier Goureau5, Christelle Monville1,2 1U861, I-Stem, Association Française contre les Myopathies (AFM), Institut National de la Santé et de la Recherche Médicale (INSERM), 2I-Stem, Association Française contre les Myopathies (AFM), Centre pour L’Etude des Cellules Souches (CECS), 4Banque de tissus humain, Hôpital Saint Louis, Assistance Publique - Hôpitaux de Paris (AP-HP), 5Sorbonne Université, INSERM, CNRS, Institut de la Vision, F-75012 We describe a method to engineer a retinal tissue composed of retinal pigment epithelial cells derived from human pluripotent stem cells cultured on top of human amniotic membranes and its preparation for grafting in animal models. Immunology and Infection Analysis of Microglia and Monocyte-derived Macrophages from the Central Nervous System by Flow Cytometry Elodie Martin1,2,3, Mohamed El-Behi1,2,3, Bertrand Fontaine1,2,3,4, Cecile Delarasse1,2,3 1Inserm U 1227, CNRS UMR 7225, 2Sorbonne Universités, UPMC, University of Paris, 3Institut du Cerveau et de la Moelle épinière, ICM, 4AP-HP, Hôpital de la Pitié Salpêtrière; This protocol provides an analysis of the macrophage subpopulations in the adult mouse central nervous system by flow cytometry and is helpful for the study of multiple markers expressed by these cells. Medicine Continuous Manual Exchange Transfusion for Patients with Sickle Cell Disease: An Efficient Method to Avoid Iron Overload Bérengère Koehl1, Florence Missud1, Laurent Holvoet1, Ghislaine Ithier1, Oliver Sakalian-Black2, Zinedine Haouari1, Emmanuelle Lesprit3, André Baruchel4, Malika Benkerrou1 1Reference Centre of Sickle Cell Disease, Hematology Unit, Robert Debré Hospital, AP-HP, 2School of Pharmacy, Université Paris Descartes, 3Etablissement Français du Sang, Robert Debré Hospital, AP-HP, 4Hematology Unit,, Robert Debré Hospital, AP-HP, Univsité Paris Diderot We have outlined a method of continuous manual exchange transfusion for the treatment of sickle cell disease in patients. This safe protocol was designed to effectively limit iron overload in patients in need of chronic transfusions and can be used extensively without any special equipment.