We explored a fluorescent strategy for sensing ochratoxin A (OTA) by using a single fluorophore-labeled aptamer for detection of OTA. This method relied on the change of the fluorescence intensity of the labeled dye induced by the specific binding of the fluorescent aptamer to OTA. Different fluorescein labeling sites of aptamers were screened, including the internal thymine bases, 3'-end, and 5'-end of the aptamer, and the effect of the labeling on the aptamer affinity was investigated. Some fluorophore-labeled aptamers showed a signal-on or signal-off response. With the fluorescent aptamer switch, simple, rapid, and selective sensing of OTA at nanomolar concentrations was achieved. OTA spiked in diluted red wine could be detected, showing the feasibility of the fluorescent aptamer for a complex matrix. This method shows potential for designing aptamer sensors for other targets.