Superoxide dismutase (SOD), a vital antioxidant enzyme ubiquitous in oxygen-metabolizing cells, plays a pivotal role in detoxifying the cellular environment. To comprehend antioxidant mechanisms fully, it is of profound importance to understand the modulation of SOD activity under various external influences, including different frequencies of low-intensity radiofrequency (RF) radiation. SOD catalyzes the conversion of superoxide free radicals into oxygen and hydrogen peroxide, contributing to the regulation of cellular oxidative stress. In this study, human cancer cells (HT 1080) were exposed to weak RF fields within the 3–5 MHz range for a duration of four days. Helmholtz coils were used to ensure controlled exposure conditions. The aim was to investigate the response of SOD activity to RF field frequencies, shedding light on potential alterations in the antioxidant defense mechanisms.