Background: Post-Operative Surgical Site Infection is a one of the most common healthcare-associated infection which has a great impact on patient safety. Knowledge of nurses play a crucial role for the prevention of spread of infection. To provide quality of care, it is very important for nurses to have good knowledge towards prevention of Post-Operative Surgical Site Infection. Aim: The study aimed to assess the Nurses' Knowledge Regarding Prevention of Post-Operative Surgical Site Infection. Design: A descriptive cross-sectional study was conducted.  Setting: Surgical wards at AL-Hilla teaching hospitals. Sample: All nursing working in surgical ward was included in study (N=50) Nurses. Tools of data collection: Data were collected through a questionnaire and self-report technique. The questionnaire includes a total of ten items of socio-demographic characteristics,28 knowledge items. Results: The mean of Nurse's knowledge regarding prevention of post-operative levels among nurses was (21.16), the majority of the study sample (65.6%) a high level of nurse's knowledge. Conclusions: As a final point, there is a significant difference between Nurse's knowledge among staff nurses and their socio-demographic characteristics. Finally; there are no significant between nurse's knowledge and (work experience in surgical ward, training regarding infection control, training program and surgical training courses). Recommendation: Education and training program should be conducted to improve nurses' knowledge and practice in some areas using evidence-based practice.