We present an integrated droplet-on-demand microfluidic platform for dispensing, mixing, incubating, extracting and analyzing by mass spectrometry pico- to nanoliter-sized droplets. All of the functional components are successfully integrated for the first time into a monolithic microdevice. Droplet generation is accomplished using computer-controlled pneumatic valves. Controlled actuation of valves for different aqueous streams enables accurate dosing and rapid mixing of reagents within droplets in either the droplet generation area or in a region of widening channel cross-section. Following incubation, which takes place as droplets travel in the oil stream, the droplet contents are extracted to an aqueous channel for subsequent ionization at an integrated nanoelectrospray emitter. Using the integrated platform, rapid enzymatic digestions of a model protein were carried out in droplets and detected on-line by nanoelectrospray ionization mass spectrometry.