Background: In 2011, the Therapeutic Goods Administration approved the use of alcohol-based rubs for hand antisepsis in surgical settings in Australia. The purpose of this research was to assess the knowledge, attitudes and perceptions of operating room healthcare professionals with respect to alcohol-based surgical rubs in order to gain insight into the potential barriers and enablers to their uptake. Methods: A descriptive, two-phase study assessed operating room healthcare professionals' knowledge, attitudes, and perceptions towards the use of alcohol-based surgical rubs within a large multi-campus public health care service in Melbourne, Australia. Results: Central to the successful uptake of alcohol-based surgical rubs is the provision of education that emphasises the significant benefits the rubs bring to patient safety and to staff occupational health, specifically with respect to hand skin integrity. Other enablers included distributing international evidence demonstrating their safety and efficacy and the revision to the Australian College of Operating Room Nurses Standard's other relevant policies, standards, and guidelines within health services. Inclusion of alcohol-based surgical rub protocols in national standards was deemed and important motivator. While most participants welcomed the introduction of alcohol-based surgical rubs, changing tradition and culture were reported as significant barriers to its uptake. Conclusions: Identifying the likely motivators and barriers to change will assist in the uptake of alcohol-based surgical rubs. Further research should examine the actual barriers and enablers to the uptake of alcohol-based surgical rubs that materialised in organisations following the approval and their introduction into Australian surgical settings.