The enzyme-bound copper of dopamine beta-monooxygenase reacted rapidly with the chelator bathocuproine disulfonate; the reaction in the presence of ascorbate was completed in 2 min at 25 degrees C with 1mM chelator. This reaction and also the reaction with EDTA could be used to prepare the apoenzyme, which in both cases was completely reactivated in less than 10 s. The reactivation data gave apparent Michaelis constants for copper 0.03 -- 0.2 micron. Trace amounts of copper in buffers and assay mixtures gave significant reactivation without added copper, unless they had been treated with a chelating resin. Titrations using the different chelation rates of free and enzyme-bound copper indicated that four copper atoms are bound per enzyme molecule of four subunits. The native enzyme was more stable against thermal inactivation than the apoenzyme, but this stability was only partially restored by addition of copper to the apoenzyme.