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Globally, Autologous blood transfusion (ABT) programs are not well developed and reasons differ from differentcountries. In East Africa including Uganda, Autologous transfusion has not exerted a significant impact on thetransfusion requirements in our Centres and there is paucity of data regarding ABT. The main objective of thisstudy was to assess knowledge and attitude of autologous blood transfusion among health workers in Fort portalregional referral hospital. Institution based cross sectional study design was used and simple random sampling wasused to get 50 health workers. Data was collected by a self-administered questionnaire and analyzed using SPSSsoftware version 21.00. More than half of the participants 30(60%) were diploma holders and majority 28(56%) hadworked for 4-6 years. 42 (84%) had ever heard of autologous blood transfusion but none had ever practiced ABT.Reasons for not practicing were lack of equipment and fear of side effects. The study found out that ABT was notbeing practiced although all the respondents were willing to practice it if introduced by the hospital. Thegovernment through ministry of health should design guidelines concerning ABT and train health workers throughCMEs