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Background: Most pharmacists have not received formal training or education in the provision of care for transgender patients. Nonetheless, pharmacists have the potential to be valuable partners in the care of transgender patients, and a continuing education course might be valuable in addressing this knowledge gap., Objective: The aim of this study was to examine the impact of a three-hour continuing education course in improving the knowledge of pharmacists to provide pharmaceutical care for transgender patients., Methods: A quasi-experimental, one-group pre-test/post-test study design was used to measure the impact of a three-hour continuing pharmacy education course on the knowledge of pharmacists on transgender care. The course was divided into three units: (1) Transgender Patient Care Introduction, (2) General Health Issues of Transgender Patients, and (3) Gender Affirming Hormone Therapy. A total of 68 pharmacists participated in the study, of which 54 completed both the pre- and post-test. An ANOVA was used to compare differences in knowledge in the group before and after the educational intervention., Results: The majority of the participating pharmacists were cisgender, heterosexual women who had not received any formal training related to transgender care. Participants demonstrated the largest increase in execution score in the third unit, with a percent improvement of 25.22% (pre-test 45.06%, post-test 70.28%; p<0.001). The average total execution score was 52.15% in the pre-test and 72.89% (p< 0.001) in the post-test., Conclusions: Pharmacists benefited from a three-hour continuing education course with an increase in knowledge regarding transgender patient care and hormone therapy for gender affirmation. As this study only evaluated the effect in short term memory, further studies are needed to assess long term impact of the continuing education course on transgender care knowledge., Competing Interests: CONFLICT OF INTEREST The authors have no affiliation or significant financial involvement in any organizations or entity with a direct financial interest in the materials or subject matter discussed in the manuscript to disclose., (Copyright: © Pharmacy Practice and the Authors.)