Although medical professionalism is a fundamental aspect of competence in medicine and a distinct facet of physicians' competence, evidence suggests that the subject of professionalism is not taught or assessed as part of medical students' curricula in Iran and many other countries. Assessing the knowledge of medical students and physicians about medical professionalism seems to be helpful in identifying the weaknesses of training in the field of professionalism and devise plans for future training on the subject.The present cross-sectional, quantitative, observational, and prevalence study recruited 149 medical interns, clinical residents, physicians, and professors working in hospitals selected through stratified random sampling using a questionnaire designed by the researchers and confirmed for its validity and reliability. The results were analyzed by Stata at a significance level of 0.05.Out of 149 cases, 61.64% were male with the mean age of 30.81 years. A total of 66 participants (44.29%) (95% confidence interval [CI]: 36.44%-52.44%) had heard and 83 (55.70%) (95% CI: 47.55%-63.55%) had not heard the term "medical professionalism" before the study. After adjusting for potential confounders, age and degree did not have statistically significant difference in assessed knowledge of medical professionalism, but sex had (mean difference: 5.88, P = 0.045), and the mean of the female was significantly higher than that of the male participants. The mean percentage of correct answers was 47.67.The present study demonstrated that the medical professionals working in the national healthcare system have an unfavorable theoretical knowledge about medical professionalism in Iran; although this does not indicate that their practices are unethical, it should be noted that one of the prerequisites of possessing a high level of medical professionalism and for establishing a proper relationship between the medical community and the patients is to have a proper knowledge of this concept. Improving behaviors and performances in medical professions requires adequate training on the concepts of medical professionalism and consequently the assessment of the levels of professionalism achieved in medical professionals.