Paper-based devices are biodegradable and have been used in diagnosis and environmental analysis field. In this work, a wax-printed paper-based analytical device combined with silver nanoparticles (AgNPs) catalyzed luminol chemiluminescence (CL) system for the determination of ofloxacin (OFLX) was presented. It was based on the enhancement of CL intensity of luminol-H2O2-OFLX system by AgNPs. Wax-printing fabrication technique was used to make the simple circle shaped paper device and large scale chips can be fabricated at the same time. Under the selected experimental conditions, a linear relationship was obtained between the CL intensity and the concentration of OFLX in the range from 1.0×10(-9) g/mL to 1.0×10(-6) g/mL with a detection limit of 3.0×10(-10) g/mL. This method has been successfully applied to the determination of OFLX in eyedrop samples.