Healthcare-Associated Infections (HAIs) are still a significant concern in healthcare facilities due to the elevated risk of transmission and their consequences for patient safety. This indicates that efforts to prevent HAIs still need to be optimized to achieve better outcomes. This study was conducted to analyze the association between knowledge, attitudes, and perceptions and HAI prevention behavior among nurses at Ratu Zalecha Martapura Hospital. The research applied a quantitative cross-sectional design involving 66 nurses selected using proportional sampling. Data were obtained through valid and reliable questionnaires, then analyzed using the chi-square test with a 95% confidence level. The findings indicated that knowledge and attitudes were significantly associated with HAI prevention behavior (p = 0.000), whereas perceptions showed no significant association with HAI prevention behavior (p = 0.061). HAI prevention behavior among nurses is associated with levels of knowledge and attitudes, but not with perceptions. Adequate knowledge and positive attitudes contribute to the development of optimal preventive behavior, highlighting the importance of continuous education and training to enhance patient safety and the quality of nursing care