Modern operating microscopes offer high power illumination to ensure optimal visualization, but can also cause thermal damage. The aim of our study is to quantify the thermal effects in vivo and discuss conditions for safe use. In a pilot study on volunteers, we measured the temperature at the skin surface during microscope illumination, including the influence of anaesthesia and the effects of staining, draping, or moistening of the skin. Irradiation within the limit given by safety regulations (200 mW/cm2) results in skin surface temperature of 43 °C. Higher intensities (forearm 335 mW/cm2, back 250 mW/cm2) are tolerated, resulting in reversible hyperaemia. At a very high illumination intensity (750 mW/cm2), pain occurs within 30 s at temperatures of 46 °C±1 °C (hand and forearm), and 43 °C±2 °C (back), respectively. Anaesthesia has no distinct effect on the temperature, whereas staining and drapes result in much higher temperatures (<100 °C). Moistening at practicable flow rates can reduce temperature efficiently when combined with a light absorbing and water absorbent drape. In conclusion, surgeons must be aware that surgical microscope illumination without protective means can cause skin temperatures to rise much above pain threshold, which in our study serves as a (conservative) benchmark for potential damage.