Many cancer cells over-express folate receptors, and this provides an opportunity for both folate-targeted fluorescence imaging and the development of targeted anti-cancer drugs. We present an optical imaging modality that allows for the monitoring and evaluation of drug delivery and release through disulfide bond reduction inside a tumor in vivo for the first time. A near-infrared folate-targeting fluorophore pair was synthesized and used to image a xenograft tumor grown from KB cells in a live mouse. The in vivo results are shown to be in agreement with previous in vitro studies, confirming the validity and feasibility of our method as an effective tool for preclinical studies in drug development.