Background: A central feature of clinical simulation training is human factors skills, providing staff with the social and cognitive skills to cope with demanding clinical situations. Although these skills are critical to safe patient care, assessing their learning is challenging. This study aimed to develop, pilot and evaluate a valid and reliable structured instrument to assess human factors skills, which can be used pre- and post-simulation training, and is relevant across a range of healthcare professions., Method: Through consultation with a multi-professional expert group, we developed and piloted a 39-item survey with 272 healthcare professionals attending training courses across two large simulation centres in London, one specialising in acute care and one in mental health, both serving healthcare professionals working across acute and community settings. Following psychometric evaluation, the final 12-item instrument was evaluated with a second sample of 711 trainees., Results: Exploratory factor analysis revealed a 12-item, one-factor solution with good internal consistency (α=0.92). The instrument had discriminant validity, with newly qualified trainees scoring significantly lower than experienced trainees ( t (98)=4.88, p<0.001) and was sensitive to change following training in acute and mental health settings, across professional groups (p<0.001). Confirmatory factor analysis revealed an adequate model fit (RMSEA=0.066)., Conclusion: The Human Factors Skills for Healthcare Instrument provides a reliable and valid method of assessing trainees' human factors skills self-efficacy across acute and mental health settings. This instrument has the potential to improve the assessment and evaluation of human factors skills learning in both uniprofessional and interprofessional clinical simulation training., Competing Interests: Competing interests: None declared.