Guidelines for the use of flow cytometry and cell sorting in immunological studies

Andrea Cossarizza (1) , Hyun-Dong Chang (2) , Andreas Radbruch (3) , Immanuel Andrä , Francesco Annunziato (4) , Petra Bacher , Regina Barnaba , Luca Battistini (5) , Wolfgang Bauer (6) , Sabine Baumgart (7) , Burkhard Becher , Wolfgang Beisker (8) , Claudia Berek (9) , Alfonso Blanco , Giovanna Borsellino (10) , David Boulais , Ryan R Brinkman (11) , Martin Büscher , Dirk Busch , Timothy P Bushnell , Xuetao Cao (12) , Andrea Cavani , Pratip K Chattopadhyay , Qingyu Cheng , Sue Chow , Mario Clerici (13) , Anne Cooke , Antonio Cosma (14) , Lorenzo Cosmi , van Duc Dang , Derek Davies , Mor de Biasi , Genny del Zotto , Silvia Della Bella , Paolo Dellabona (15) , Günnur Deniz , Mark Dessing , Andreas Diefenbach , James P. Di Santo (16) , Francesco Dieli , Bo Dolf , Vera S Donnenberg , Thomas Dörner (17) , Götz R A Ehrhardt , Bo Endl , Pablo Engel , Britta Engelhardt , Charlotte Esser , Bart Everts , Christine Falk , Todd A Fehniger , Andrew Filby , Simon Simon , Marie Follo , Bo Förster , John Foster , John Foulds , Paul S Frenette , Ari Galbraith , Bo Garbi , Maria Dolores García-Godoy , Kamran Ghoreschi , Mor Gibellini , Christoph Goettlinger , Carl S Goodyear , Andrea Gori (18) , Francisco Grogan , Mor Gross , Andreas Grützkau (19) , Daryl Grummitt , Alexander Hahn , Quirin Hammer , Anja E Hauser , David Houston , David Hedley , Guadalupe Herrera , Alexander Herrmann (20) , Falk Hiepe , Bo Holland , Pleun Hombrink , Jessica P Houston , Bimba F Hoyer , Ton Huang , Christopher A Hunter , Anna Iannone , Hans-Martin Jäck , Beatriz Jávega , Stipan Jonjic (21) , Kerstin Juelke , Steffen Jung (22) , Toralf Kaiser , Tomas Kalina , Baerbel Keller , Srijit Khan , Alexander Kienhöfer , Thomas Kroneis , Désirée Kunkel (23) , Christian Kurts (24) , Pia Kvistborg , Virginia Lannigan , Olivier Lantz (25) , Anis A Larbi (26) , Salomé Leibundgut-Landmann (27) , Michael D Leipold , Megan K Levings , Virginia Litwin , Yanling Liu (28) , Michael Lohoff (29) , Giovanna Lombardi (30) , Lilly Lopez , Amy Lovett-Racke , Erik Lubberts (31) , Burkhard Ludewig , Enrico Lugli (32) , Holden T Maecker , Gloria Martrus (33) , Giuseppe Matarese , Alexander Maueröder , Mairi Mcgrath (34) , Iain Mcinnes , Henrik E Mei , Fritz Melchers , Susanne Melzer , Dirk Mielenz , Kingston Mills , Jenny Mjösberg (35) , Jonni Moore , Barry Moran , Alessandro Moretta (36) , Lorenzo Moretta (36) , David Mosmann , Susann Müller (37) , Werner Müller (38) , Christian Münz (39) , Gabriele Multhoff (40) , Alexander Munoz , Kenneth M Murphy , Toshinori Nakayama , Milena Nasi , Christine Neudörfl , John Nolan (41) , Sussan Nourshargh (42) , José-Enrique O'Connor , Francisco Ouyang , Annette Oxenius , Raghav Palankar , Isabel Panse (2) , Peter Peterson , Christian Peth , Jordi Petriz , Daisy Philips , Winfried Pickl , Regina Piconese , Marcello Pinti , a Graham Pockley , Alexander Podolska , Carlo Pucillo , David Quataert , Timothy R D J Radstake , Bartek Rajwa (43) , David Rebhahn , Diether Recktenwald , Ester B M Remmerswaal , Katy Rezvani , Laura G Rico , J Paul Robinson , Chiara Romagnani , Anna Rubartelli , Jürgen Ruland , Shimon Sakaguchi (44) , Francisco Sala-De-Oyanguren , Yvonne Samstag , Sharon Sanderson , Birgit Sawitzki , Alexander Scheffold , Matthias Schiemann , Frank Schildberg , Esther Schimisky , Stephan A Schmid , Steffen Schmitt (45) , Kilian Schober , Thomas Schüler (46) , Axel Ronald Schulz , Ton Schumacher , Cristiano Scotta , T Vincent Shankey , Anat Shemer , Anna-Katharina Simon , Josef Spidlen (11) , Alan M Stall , Regina Stark , Christina Stehle , Merle Stein , Tobit Steinmetz , Hannes Stockinger (47) , Yousuke Takahama , Attila Tarnok (48) , Zhigang Tian (49) , Gergely Toldi , Julia Tornack , Elisabetta Traggiai , Joe Trotter , Paulo Ulrich , Marlous van Der Braber , René a W van Lier , Marcello Veldhoen , Salvador Vento-Asturias , Paulo Vieira (50) , David Voehringer (51) , Hans-Dieter Volk (3) , Konrad von Volkmann , Ari Waisman (52) , Rachael Walker , Michael D Ward , Klaus Warnatz , Sarah Warth , James Watson , Carsten Watzl , Leonie Wegener , Annika Wiedemann , Jürgen Wienands , Gerald Willimsky , James Wing , Peter Wurst , Liping Yu , Alice Yue , Qianjun Zhang , Yi Zhao (53) , Susanne Ziegler (54) , Jakob Zimmermann

1 Dipartimento di Scienze Biomediche
2 Cell Biology
3 Klinik für Dermatologie, Venerologie und Allergologie
4 Department of Internal Medicine
5 Neuroimmunology Unit
6 Inorganic Chemistry II
7 Caprotec Bioanalytics GmbH
8 IOTA ES - International Occultation Timing Association European Section
9 Institut der Leibniz-Gemeinschaft, Berlin
10 IRCCS - Fondazione Santa Lucia
11 Terry Fox Laboratory
12 Department of Immunology
13 Fondazione Don Carlo Gnocchi
14 IMVA - U1184 - Immunologie des Maladies Virales et Autoimmunes
15 Experimental Immunology Unit, Dept. of Oncology
16 Département d'Immunologie - Department of Immunology
17 Charité Hospital
18 UB - Universitat de Barcelona
19 Rheumatologie
20 Department of Internal Medicine I
21 Department of Histology and Embryology
22 Weizmann Institute of Science [Rehovot, Israël]
23 Régulation des Infections Rétrovirales
24 Institutes of Molecular Medicine and Experimental Immunology
25 U932 - Immunité et cancer
26 SIgN - Singapore Immunology Network
27 Institute of Virology [Zürich]
28 College of Food Science and Technology [Shangai]
29 Institute for Medical Microbiology and Hygiene
30 MRC - Centre for Transplantation, King's College London
31 Erasmus MC - Erasmus University Medical Center [Rotterdam]
32 Unit of Clinical and Experimental Immunology
33 Heinrich Pette Institute [Hamburg]
34 Institute of Translational Medicine
35 Department of Medicine Huddinge
36 UniGe - Università degli studi di Genova = University of Genoa
37 Department of Environmental Microbiology
38 Experimental Immunology
39 Viral Immunobiology
40 Department of Radiation Oncology [Munich]
41 Department of Mathematics and Statistics
42 William Harvey Research Institute, Barts and the London Medical School
43 Cytometry Laboratories and School of Veterinary Medicine
44 Osaka University [Osaka]
45 FACS and Array Core Facility
46 Institute for Cognitive Science
47 Department of Molecular Immunology
48 Universität Leipzig [Leipzig]
49 Institute of Immunology
50 Lymphopoïèse (UMR_1223 / U1223 / U-Pasteur_4) - Lymphopoïèse
51 Institute for Immunology
52 Department of Internal Medicine
53 PROTEE - Processus de Transfert et d'Echanges dans l'Environnement - EA 3819
54 Heinrich-Pette-Institut

The classical model of hematopoiesis established in the mouse postulates that lymphoid cells originate from a founder population of common lymphoid progenitors. Here, using a modeling approach in humanized mice, we showed that human lymphoid development stemmed from distinct populations of CD127(-) and CD127(+) early lymphoid progenitors (ELPs). Combining molecular analyses with in vitro and in vivo functional assays, we demonstrated that CD127(-) and CD127(+) ELPs emerged independently from lympho-mono-dendritic progenitors, responded differently to Notch1 signals, underwent divergent modes of lineage restriction, and displayed both common and specific differentiation potentials. Whereas CD127(-) ELPs comprised precursors of T cells, marginal zone B cells, and natural killer (NK) and innate lymphoid cells (ILCs), CD127(+) ELPs supported production of all NK cell, ILC, and B cell populations but lacked T potential. On the basis of these results, we propose a "two-family" model of human lymphoid development that differs from the prevailing model of hematopoiesis.

Keywords: natural killer and innate lymphoid cells differentiation; early lymphoid progenitors; B cell ontogeny; Human lymphopoiesis; T cell precursors

Domains: Life Sciences [q-bio]; Immunology

Andrea Cossarizza, Hyun-Dong Chang, Andreas Radbruch, Immanuel Andrä, Francesco Annunziato, et al.. Guidelines for the use of flow cytometry and cell sorting in immunological studies. European Journal of Immunology, 2017, 47 (10), pp.1584 - 1797. ⟨10.1002/eji.201646632⟩. ⟨pasteur-01619848⟩