Importance: A lack of health care provider knowledge and training has been identified as one factor that contributes to health disparities for sexual and gender minority (SGM) populations. Objective: To explore occupational therapy practitioners’ self-reported knowledge about, clinical preparedness for, and attitudes toward working with lesbian, gay, bisexual, and transgender (LGBT) clients. Design: Online survey of occupational therapy practitioners. Participants and Setting: Respondents were recruited by means of snowball sampling through social media groups, state occupational therapy association websites, and emails. Surveys were posted to electronic occupational therapy social media sites. Measures: Knowledge, clinical preparedness, and attitudes were measured using the Lesbian, Gay, Bisexual, and Transgender Development of Clinical Skills Scale (LGBT–DOCSS). Information about demographic characteristics, education, and workplace was also collected. Results: Five hundred eighty-nine occupational therapy practitioners responded to the survey. Degree level (master’s vs. bachelor’s or less), continuing education (minimum 1–2 hr of LGBT-specific training), practice setting (mental health), minority sexual orientation, and having a close friend or family member who identifies as SGM were associated with higher mean scores on the LGBT–DOCSS. Higher religiosity and frequency of religious practice were associated with lower scores on knowledge and attitudinal awareness. Conclusions and Relevance: Occupational therapy practitioners often care for clients from backgrounds and cultures that differ from their own. Identifying gaps in education and opportunities for fostering LGBT-positive attitudes can facilitate the development of programs to improve practice with LGBT clients and help measure the effectiveness of such programs. What This Article Adds: This study provides evidence that a basic level of continuing education can improve occupational therapy practitioners’ knowledge of and skills for working with LGBT populations and highlights the need to examine and change structural biases.