In all biological systems, protein amount is a function of the rate of production and clearance. The speed of a response to a disturbance in protein homeostasis is determined by turnover rate. Quantifying alterations in protein synthesis and clearance rates is vital to understanding disease pathogenesis (e.g., aging, inflammation). No methods currently exist for quantifying production and clearance rates of low-abundance (femtomole) proteins in vivo. We describe a novel, mass spectrometry—based method for quantitating low-abundance protein synthesis and clearance rates in vitro and in vivo in animals and humans. The utility of this method is demonstrated with amyloid-β (Aβ), an important low-abundance protein involved in Alzheimer’s disease pathogenesis. We used in vivo stable isotope labeling, immunoprecipitation of Aβ from cerebrospinal fluid, and quantitative liquid chromatography electrospray-ionization tandem mass spectrometry (LC-ESI-tandem MS) to quantify human Aβ protein production and clearance rates. The method is sensitive and specific for stable isotope-labeled amino acid incorporation into CNS Aβ (+1% accuracy). This in vivo method can be used to identify pathophysiologic changes in protein metabolism and may serve as a biomarker for monitoring disease risk, progression, or response to novel therapeutic agents. The technique is adaptable to other macromolecules, such as carbohydrates or lipids.