biostudies-literatureHagberg CEKarolinska Institutet/AstraZeneca Integrated Cardio Metabolic Centre (KI-AZ ICMC)Novo Nordisk FoundationThe Erling-Persson Family FoundationEuropean Research CouncilVallee FoundationWilhelm och Else Stockmanns StiftelseKarolinska InstitutetSwedish Research CouncilDiabetes Research Program at Karolinska InstitutetNovo Nordisk FondenSwedish Society for Medical Research2746-2756.e5https://www.ebi.ac.uk/biostudies/studies/S-EPMC6137819biostudies-literatureUnknown24(10)Adipocytes, once considered simple lipid-storing cells, are rapidly emerging as complex cells with many biologically diverse functions. A powerful high-throughput method for analyzing single cells is flow cytometry. Several groups have attempted to analyze and sort freshly isolated adipocytes; however, using an adipocyte-specific reporter mouse, we demonstrate that these studies fail to detect the majority of white adipocytes. We define critical settings required for adipocyte flow cytometry and provide a rigid strategy for analyzing and sorting white and brown adipocyte populations. The applicability of our protocol is shown by sorting mouse adipocytes based on size or UCP1 expression and demonstrating that a subset of human adipocytes lacks the β₂-adrenergic receptor, particularly in the insulin-resistant state. In conclusion, the present study confers key technological insights for analyzing and sorting mature adipocytes, opening up numerous downstream research applications.Cell reportsFlow Cytometry of Mouse and Human Adipocytes for the Analysis of Browning and Cellular Heterogeneity.PMC6137819261258NNF12OC1016064Shabalina IGThorell ALi QKozina VBhowmick DNedergaard JKutschke MHarms MJShilkova OSpalding KLKiss EHagberg CEBoucher JfalseFlow Cytometry of Mouse and Human Adipocytes for the Analysis of Browning and Cellular Heterogeneity.Adipocytes, once considered simple lipid-storing cells, are rapidly emerging as complex cells with many biologically diverse functions. A powerful high-throughput method for analyzing single cells is flow cytometry. Several groups have attempted to analyze and sort freshly isolated adipocytes; however, using an adipocyte-specific reporter mouse, we demonstrate that these studies fail to detect the majority of white adipocytes. We define critical settings required for adipocyte flow cytometry and provide a rigid strategy for analyzing and sorting white and brown adipocyte populations. The applicability of our protocol is shown by sorting mouse adipocytes based on size or UCP1 expression and demonstrating that a subset of human adipocytes lacks the β₂-adrenergic receptor, particularly in the insulin-resistant state. In conclusion, the present study confers key technological insights for analyzing and sorting mature adipocytes, opening up numerous downstream research applications.2018-01-01T00:00:00Z2018 Sep2026-06-08T04:04:50.495Z2019-03-26T23:56:40ZS-EPMC61378193018450710.1016/j.celrep.2018.08.006