ARTICLE Single cell RNA sequencing of human microglia uncovers a subset associated with Alzheimer’s disease Marta Olah 1,2,3,4,19 , Vilas Menon 1,2,3,4,19 , Naomi Habib 4,5 , Mariko F. Taga 1,2,3,4 , Yiyi Ma 1,2,3,4 , Christina J. Yung 1 , Maria Cimpean 1 , Anthony Khairallah 1 , Guillermo Coronas-Samano 2,6 , Roman Sankowski 7,8 , Dominic Grün 9 , Alexandra A. Kroshilina 1 , Danielle Dionne 4 , Rani A. Sarkis 10 , Garth R. Cosgrove 11 , Jeffrey Helgager 12 , Jeffrey A. Golden 12 , Page B. Pennell 10 , Marco Prinz 7,13,14 , Jean Paul G. Vonsattel 2,6 , Andrew F. Teich 2,3,6 , Julie A. Schneider 15 , David A. Bennett 15 , Aviv Regev 4,16,17,18 , Wassim Elyaman 2,3,4 , Elizabeth M. Bradshaw 2,3,4 & Philip L. De Jager 1,2,3,4 ✉ The extent of microglial heterogeneity in humans remains a central yet poorly explored question in light of the development of therapies targeting this cell type. Here, we investigate the population structure of live microglia purified from human cerebral cortex samples obtained at autopsy and during neurosurgical procedures. Using single cell RNA sequencing, we find that some subsets are enriched for disease-related genes and RNA signatures. We confirm the presence of four of these microglial subpopulations histologically and illustrate the utility of our data by characterizing further microglial cluster 7, enriched for genes depleted in the cortex of individuals with Alzheimer’s disease (AD). Histologically, these cluster 7 microglia are reduced in frequency in AD tissue, and we validate this observation in an independent set of single nucleus data. Thus, our live human microglia identify a range of subtypes, and we prioritize one of these as being altered in AD. https://doi.org/10.1038/s41467-020-19737-2 OPEN 1 Center for Translational and Computational Neuroimmunology, Columbia University Medical Center, New York, NY, USA. 2 Taub Institute for Research on Alzheimer’s Disease and Aging Brain, Columbia University Medical Center, New York, NY, USA. 3 Department of Neurology, Columbia University Medical Center, New York, NY, USA. 4 Cell Circuits Program, Broad Institute, Cambridge, MA, USA. 5 Edmond & Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel. 6 Department of Pathology and Cell Biology, Columbia University Medical Center, New York, NY, USA. 7 Institute of Neuropathology, Medical Faculty, University of Freiburg, Freiburg, Germany. 8 Berta-Ottenstein-Programme for Clinician Scientists, Faculty of Medicine, University of Freiburg, Freiburg, Germany. 9 Max-Planck-Institute of Immunobiology and Epigenetics, Freiburg, Germany. 10 Department of Neurology, Brigham and Women’s Hospital, Boston, MA, USA. 11 Department of Neurosurgery, Brigham and Women’s Hospital, Boston, MA, USA. 12 Department of Pathology, Brigham and Women’s Hospital, Boston, MA, USA. 13 Signaling Research Centers BIOSS and CIBSS, University of Freiburg, Freiburg, Germany. 14 Center for NeuroModulation, Faculty of Medicine, University of Freiburg, Freiburg, Germany. 15 Rush Alzheimer’s Disease Center, Rush University Medical Center, Chicago, IL, USA. 16 Klarman Cell Observatory, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA. 17 Howard Hughes Medical Institute, Department of Biology, MIT, Cambridge, MA 02140, USA. 18 Present address: Genentech, 1 DNA Way, South San Francisco, CA 94080, USA. 19 These authors contributed equally: Marta Olah, Vilas Menon. ✉ email: pld2115@cumc.columbia.edu NATURE COMMUNICATIONS | (2020)11:6129 | https://doi.org/10.1038/s41467-020-19737-2 | www.nature.com/naturecommunications 1 1234567890():,;