Abstract: The specific absorption rate (SAR) and temperature changes in MC3TC-E1 cells layer exposed to 2.45 GHz Wi-Fi irradiation in close range (3 cm) were simulated by finite-difference time-domain (FDTD)method. The cells reactive oxygen species (ROS) was detected. The energy absorption and heating of 2.45 GHz radio frequency radiation on osteoblasts in vitro, and its non-thermal effects on ROS in osteoblasts were analyzed. The results show that the average cell temperature is increased by 0.5℃ and the highest temperature is increased by less than 1℃ after 90-minute Wi-Fi irradiation at 500 mW 2.45 GHz; the average SAR value of cell culture dish is 0.835 6 W/kg, the maximum value is 1.259 4 W/kg, and the minimum value is 0.553 7 W/kg; Wi-Fi radiation increases cell ROS; ROS and SAR are positively correlated; the temperature in the sham group with increasing temperature of 1℃ does not affect ROS statistically; the non-thermal effect of 2.45 GHz radio frequency induces the increase of ROS in osteoblasts.