The electrochemical, aptamer-based (E-AB) sensor platform provides a modular approach to the continuous, real-time measurement of specific molecular targets (irrespective of their chemical reactivity) in situ in the living body. To achieve this, however, requires the fabrication of sensors small enough to insert into a vein, which, for the rat animal model we employ, entails devices less than 200 μm in diameter. The limited surface area of these small devices leads, in turn, to low faradaic currents and poor signal-to-noise ratios when deployed in the complex, fluctuating environments found in vivo. In response we have developed an electrochemical roughening approach that enhances the signaling of small electrochemical sensors by increasing the microscopic surface area of gold electrodes, allowing in this case more redox-reporter-modified aptamers to be packed onto the surface, thus producing significantly improved signal-to-noise ratios. Unlike previous approaches to achieving microscopically rough gold surfaces, our method employs chronoamperometric pulsing in a 5 min etching process easily compatible with batch manufacturing. Using these high surface area electrodes, we demonstrate the ability of E-AB sensors to measure complete drug pharmacokinetic profiles in live rats with precision of better than 10% in the determination of drug disposition parameters.