Introduction: Determining patients’ treatment priority based on the severity of their condition, and choosing the best treatment for the highest possible number of patients in the shortest time is called triage. Although triage has only been used in Iranian hospitals since a few years ago, almost all hospital emergency services over the country currently have this unit. Nurses are responsible for triaging patients, but there is little data on their knowledge and quality of practice in this regard. Therefore, this study was aimed to assess the knowledge level and practice of emergency department (ED) nurses regarding hospital triage. Methods: This study is a prospective cross-sectional one, carried out with the aim of evaluating the knowledge level and practice of nurses regarding hospital triage. Simple random sampling was used to select ED nurses from teaching hospitals in Yazd province. Data were gathered using a questionnaire, validity and reliability of which had been confirmed before. The questionnaire comprised of 3 parts, 11 demographic questions, 15 questions evaluating knowledge, and 10 assessing practice. Data were analyzed using SPSS 16.0 and descriptive and inferential statistical tests. Results: A total of 45 questionnaires were filled and gathered. 62.2% of cases were male with the mean age of 35.92 ± 5.74 years, and 37.8% were female with the mean age of 30.82 ± 7.37 years. Mean age of all participants was 34 ± 6.80 years. Mean work experience of the studied nurses was 10.07 ± 6.89 years and their mean work experience in ED was 5.31 ± 4.70 years. Evaluation of the second part, regarding their knowledge, revealed that 42.07% of the answers to the questions were correct and their mean score was 6.33 ± 1.96. In addition, their practice assessment showed that 50.44% of the answers given in this part were right and their mean score in this regard was 5 ± 1.47. In total, 45.33% of the answers given by the participants were correct regarding the scenarios. Mean overall score (sum of the scores obtained from knowledge and practice parts) for the participants was 11.33 ± 2.50. Statistical analysis found a significant correlation only between the working shift and the knowledge score (p = 0.003). Conclusion: It seems that the knowledge and experience of the nurses working in EDs of centers under supervision of Shahid Sadoughi University of Medical Sciences, Yazd, Iran, regarding triage, is below average. Consequently, it is suggested to hold special retraining courses in the field of triage for the staff in order to improve the quality of care.