Purpose: To evaluate the effects of increasing thermocycling regimens on bond strength and nanoleakage of an etch-and-rinse adhesive. Materials and Methods: Adper Single Bond (3M ESPE) was used for bonding resin composite to human enamel and dentin. Specimens were stored in water (37°C, 24 h) and subjected to thermocycling (5°C and 55°C) in five groups: 1. control (no thermal cycling), 2. 100, 3. 500, 4. 2000, or 5. 10,000 cycles. The microshear bond test was performed at a crosshead speed of 1 mm/min (n = 16). Nanoleakage specimens were immersed in silver nitrate and FE-SEM micrographs were digitally analyzed to calculate silver penetration in three samples. Results: The mean bond strength to enamel was not significantly affected by thermocycling. However, artificial aging decreased dentin bond strength in groups 4 and 5 compared to control. Weibull analysis indicated that the characteristic strength decreased gradually with aging in both substrates. All groups exhibited some nanoleakage, and a significant increase in silver penetration compared to the control was observed in enamel and dentin groups 2 to 5. Conclusion: A nanoleakage increase was detected in an earlier stage of aging than when a significant drop in bond strength was observed, with the dentin bond being more susceptible to deterioration. Keywords: dental adhesive, thermocycling, nanoleakage, bond strength