Introduction: Hospital-acquired infection is a major safety issue affecting the quality of care of hundreds of millions of patients every year, in both developed and developing countries, including Ethiopia. Asepsis is a set of process to prevent the spread of hospital acquired infection. To protect patients from pathogens during medical and surgical procedures, health care forces use aseptic techniques. Objectives: To assess the efficacy of hands on practice coaching regarding Operation room asepsis among nurses in Dessie Referral Hospital, Eastern Amhara, Ethiopia. Methodology: Institution based pre-experimental one group pre and post test design was used and 51 study subjects were selected by convenient sampling technique. The study samples included nurses working at Operation theatre. Data collection took place between October-November 2016. The collected data were analyzed using descriptive and inferential statistics and statistical significances for variables were set at p- value less than 0.05. Results: There was a significant difference in skill score related factors under this study before and after the hands on practice coaching. There was a significant median difference in skill (p = 0.02) scores after the hands on practice coaching regarding operation room asepsis among study subjects. Sixty-seven (67) % of respondents rated the hands on practice coaching as very good. Seventeen (17) % of respondents had rated as good, and 4% rated as fair and also 4% of the respondents rated as poor. Conclusion and Recommendation: The hands on practice coaching provided by Wollo University staff was found to be efficient, conceivable on operation room asepsis skill and which can be scaled up further to nearby Hospital staff. Periodic training program shall be provided to nurses who work at operation theatre to update their skill regarding operation room asepsis.