Purpose: This study aimed to identify the factors affecting compliance with personal protective equipment (PPE) use among medical device reprocessing staff. Methods: This descriptive cross-sectional study included 163 cleaning staff members from ten general hospitals in Seoul and Gyeonggi. Data were collected using self-report questionnaires administered between July and September 2023. Analysis included t-tests, ANOVA, Pearson"s correlation coefficient, Bonferroni correction, and multiple regression, conducted using SAS ver.9.4. Results: Statistically significant differences in compliance with PPE were found based on department and exposure to contamination within six months (t=-2.82, p=.007). Attitudes toward PPE (r=.22, p=.006) and awareness of the safety climate (r=.22, p=.006) showed a statistically significant positive correlation with PPE compliance. Factors influencing use of personal protective equipment by cleaning staff during medical device reprocessing were department, compliance with PPE, and awareness of the safety climate. The explanatory power of these factors was 58.0%. Conclusion: Improving PPE compliance and creating a safe cleaning environment entails fostering a supportive safety climate. Additionally, regular training that takes into consideration the characteristics of the cleaning staff, alongside continuous monitoring, is required.