To examine nursing graduates' knowledge of functions and limitations of pulse oximetry.Pulse oximetry is a technology ubiquitous in its use in modern clinical settings. Although the technology's ability to accurately reflect hypoxaemia in patients has been established, its contribution to improving patient outcomes is less certain. In addition, experienced nurses have previously demonstrated poor understanding of the limitations of the technology. Pregraduate education has been identified as a potential source of knowledge deficit and has been recommended by authors as an important target of investigation.Cross-sectional, comparative, multicentre study.A previously published and validated tool was used with the addition of eight clinical scenario questions which were validated by an expert panel. Convenience sampling was used to select participants to form one cohort of newly graduated nurses (Cohort 1: n = 210) and a second cohort of nurses completing their intensive postgraduate clinical year (Cohort 2: n = 97).Significant deficits relating to the theoretical factors that affect pulse oximetry application and interpretation were identified. Results suggest some knowledge is negatively correlated with clinical experience and that pregraduate university education appears to influence the ability to effectively apply pulse oximetry knowledge to clinical scenarios.This study provides insight into pulse oximetry knowledge acquisition and deficits of graduate nurses which may inform pre- and postgraduate nurse education. In particular, it suggests that some pulse oximetry knowledge and clinical application of knowledge is not enhanced by clinical experience but rather is currently gained through pregraduate experiences.Inappropriate utilisation and interpretation of pulse oximetry places patients at risk of mismanagement and undetected deterioration. Improvement in pregraduate education around the appropriate use of pulse oximetry may reduce clinical costs, reduce incidents of failure to rescue and improve patient outcomes.