Objective To explore the level and factors associated with evidence-based nursing (EBN) practice in pain assessment. Methods A total of 279 nurses were invited to complete the pain assessment evidence-based nursing practice scale. Results The score of EBN practice in pain assessment was 105.20±7.70 and the score of practicability was 108.65±11.30.Education background, working department, whether or not to build pain management rules and regulations, whether or not to treat pain as the fifth vital sign, pain training and professional title were factors influencing EBN practice in pain assessment (P＜0.05,P＜0.01). Conclusion Nurses report moderate level of EBN practice in pain assessment. Nursing managers should improve pain assessment regulations and regularly provide training regarding pain assessment to enhance quality of pain management.