Abstract Background Capturing general aspects of the occupational subculture of nursing is needed in long-term care (LTC) given it's latent influence on the quality of care that residents receive and on the ability of nursing staff (licensed nurses and certified nursing assistants) to implement evidence-based practice innovations. The psychometrically validated Nursing Culture Assessment Tool (NCAT) provides a comprehensive assessment using six dimensions (teamwork, communication, satisfaction, professional commitment, behaviors, and expectations), and evaluation of these dimensions could help positively reshape the culture before any change implementation. Aims To: (1) assess the validity and reliability of the NCAT across nursing staff in a single type of clinical setting – LTC facilities, and (2) present a refined theoretical model of the interaction of culture and practice implementation. Method A cross-sectional, exploratory investigation of the NCAT in LTC settings was conducted. Empirical construct validity of the nineteen item NCAT's six subscales was investigated by confirmatory factor analysis using a sample of licensed nurses and certified nursing assistants (n = 318). Results The model fit was judged using the comparative fit index (0.94) and standardized root mean-square residual (0.05). Cronbach's alpha correlation coefficients of items in each subscale and in the overall scale ranged from 0.76 to 0.94. Conclusion A summary of the NCAT development and report on its psychometric properties when administered in LTC settings is provided, extending previous findings of the NCAT's enhanced stability when used in assessing nursing staff perceptions in LTC and by demonstrating the NCAT is a reliable and valid psychometric screening tool for nursing culture.