Introduction The use of fluoroscopy-assisted surgical procedures has been increasing recently. The extensive use of fluoroscopy, combined with the lack of knowledge about radiation risks among operating room (OR) personnel (surgeons, anaesthetists, nurses, and radiographers), may lead to misconceptions. The perceptions and beliefs of any health professional concerning radiation protection safety (RPS) may affect their behaviour during surgery, leading to negative outcomes. The aim of this study was to construct a new, original, reliable, and valid instrument to record the beliefs, perceptions, and behaviours of health professionals during surgery, which reflects the established culture of RPS. Methods A questionnaire was designed, consisting of 95 questions grouped into six coherent sections. The study was performed at a Greek public hospital with a high workload in terms of fluoroscopically guided surgical operations. Results It was distributed among 136 people, and 132 completed questionnaires were collected (response rate 97%). Exploratory factor analysis was performed separately for each scale studied and Cronbach's Alpha reliability analysis was also performed. The reliability of the greater part of the new measurement tool ranged from very good to acceptable. Conclusion The questionnaire developed in this study is a valid and reliable option for recording health professionals' perceptions, beliefs, and behaviours concerning the RPS culture. Implications for practice This study helps clinical radiographers to understand a possible knowledge gap about RPS and locates the specific fields that the OR personnel misunderstand. It also can support and establish local learning organisations with regular targeted staff training for health professionals, thereby leading to a generally improved RPS culture.

(1) Background: Health professionals' knowledge, beliefs and perceptions concerning radiation protection may affect their behaviour during surgery and consequently influence the quality of health services. This study highlights the health professionals' average knowledge level and captures the beliefs, perceptions, and behaviours in a large public Greek hospital. (2) Materials and Methods: A cross-sectional study was carried out, including health professionals working in operating rooms. One hundred thirty-two staff members participated by responding to an original questionnaire. The sample consisted of nurses, radiographers and medical doctors of various specialties involved daily in surgical procedures where ionizing radiation is required. The survey was conducted from March to June 2021, and the response rate was 97%. (3) Results: The level of overall knowledge of health professionals regarding radiation protection safety was not satisfactory. Females and employees with a lower level of education had more misconceptions about radiation and radiation protection. Employees of younger ages and with less previous experience were more likely to have negative emotions towards radiation exposure. Finally, employees with fewer children tended to express physical complaints caused by their negative emotions due to radiation exposure. (4) Conclusions: Health professionals' lack of basic and specialized knowledge concerning radiation protection safety had a negative impact on the provision of health services. The continuing training of the staff seemed to be the only solution to reverse this trend. The training should highlight how radiation exposure can be minimized, safeguarding health professionals' trust and sense of security by significantly improving their working environment.