The effect of gamma irradiation therapy on the ultimate tensile strength (UTS) of enamel and dentin in relation to prism orientation, dentin tubule orientation, and location is unknown. It was hypothesized that tubule and prism orientation, location, and irradiation have an effect on the UTS of dental structures. Forty human third molars were used, half of which were subjected to 60 Gy of gamma irradiation, in daily increments of 2 Gy. The specimens were evaluated by microtensile testing. Results showed that irradiation treatment significantly decreased the UTS of coronal and radicular dentin and of enamel, regardless of tubule or prism orientation. With or without irradiation, enamel was significantly stronger when tested parallel to its prismatic orientation. Coronal and radicular dentin of non-irradiated specimens presented significantly higher UTS when tested perpendicularly to tubule orientation. However, when the teeth were irradiated, the influence of tubule orientation disappeared, demonstrating that irradiation is more harmful to organic components.