The use of ultraviolet fluorescence markers in medical simulations has become popular in recent years, especially during the COVID-19 pandemic. Healthcare workers use ultraviolet fluorescence markers to replace pathogens or secretions, and then calculate the regions of contamination. Health providers can use bioimage processing software to calculate the area and quantity of fluorescent dyes. However, traditional image processing software has its limitations and lacks real-time capabilities, making it more suitable for laboratory use than for clinical settings. In this study, mobile phones were used to measure areas contaminated during medical treatment. During the research process, a mobile phone camera was used to photograph the contaminated regions at an orthogonal angle. The fluorescence marker-contaminated area and photographed image area were proportionally related. The areas of contaminated regions can be calculated using this relationship. We used Android Studio software to write a mobile application to convert photos and recreate the true contaminated area. In this application, color photographs are converted into grayscale, and then into black and white binary photographs using binarization. After this process, the fluorescence-contaminated area is calculated easily. The results of our study showed that within a limited distance (50–100 cm) and with controlled ambient light, the error in the calculated contamination area was 6%. This study provides a low-cost, easy, and ready-to-use tool for healthcare workers to estimate the area of fluorescent dye regions during medical simulations. This tool can promote medical education and training on infectious disease preparation.