Background: Operating Rooms (ORs) are complicated environments that necessitate the improvement of OR staff's knowledge and skills to remain clinically competent and secure patient safety. The aim of this study was to assess clinical competence of OR staff in accordance with some related factors. Materials and Methods: This descriptive analytical cross-sectional study was conducted on 227 OR staff in nine academic hospitals. Sampling was performed from the beginning to the end of May 2019 and the samples were selected by quota sampling. Data were collected using a researcher-made questionnaire encompassing six dimensions of competency including general knowledge, specialized knowledge, general practical skills, specific practical skills, personality, and motivation. Data analysis was performed using descriptive and interpretive statistics. Results: The mean (SD) total score of competence was 80.99, which was optimal (11.28). The lowest score was related to the dimension of general practical skills with the mean (SD) score of 53.32 (10.26). The mean score of specialized practical skills was significantly higher in single-specialty ORs (F = 21.53, p < 0.001). Based on multiple linear regression test, it was possible to predict clinical competency through the age and work experience (R-squared = 0.96, beta = 0.31, p = 0.022). Conclusions: Specialized training has overshadowed the general practical skills that are related to the observation of basic principles of patient safety apart from surgical specialization. Strengthening of competence in general practical skills need to be prioritized in empowerment programs. We need a fixed and permanent space for the continuation of educational programs designed to promote perioperative general practical skills.