Updated project metadata. The turnover of brain proteins is critical for organism survival, and its perturbations are strongly linked to pathology. Nevertheless, precise protein lifetimes have been difficult to obtain in vivo. They are readily measured in vitro by feeding cells with isotopically-labeled amino acids, followed by mass spectrometry analyses. In vivo proteins are generated from at least two sources: labeled amino acids from the diet, and non-labeled amino acids from the degradation of pre-existing proteins. This renders measurements difficult. We solved this problem rigorously by developing a novel workflow that combines mouse in vivo isotopic labeling with mass spectrometry and mathematical modeling. We also established several independent approaches to test and validate the results. This enabled us to measure, for the first time, the accurate lifetimes of ~3500 brain proteins. The high precision of our data provided a large set of biologically-significant observations, including pathway-, organelle-, organ-, or cell-specific effects, along with a comprehensive catalogue of stable proteins, including novel extremely long-lived proteins (ELLPs).