Isolation, Culture, and Functional Analysis of Murine Thermogenic Adipocytes

Studying brown and brite adipose tissue requires precise and reliable quantification of cellular thermogenesis. This protocol describes the isolation of primary murine pre-adipocytes, differentiation into thermogenic brown and brite adipocytes, and subsequent oxygen consumption analysis. Commonly applied procedures only mea-sure basal and maximal proton leak-linked oxygen consumption but not explicitly un-coupling protein 1 (UCP1)-dependent respiration. Meaningful oxygen consumption analyses require (1) the activation of UCP1, (2) control over intracellular free-fatty -acid levels, and (3) inhibition of ATP-consuming futile cycles.For complete details on the use and execution of this protocol, please refer to Li et al. (2014, 2017, 2018) and Schweizer et al. (2018).