Background: Transgender people face numerous barriers to accessing care, particularly in rural settings. Transportation, travel time, a lack of providers offering transgender care, and discrimination all contribute to these barriers. The Gender Wellness Center was established in New York State, USA, to fill a gap in rural transgender care and was subsequently awarded a Robert Wood Johnson Foundation grant to establish a Center of Excellence. This study examined the implementation of the Center of Excellence, a complex intervention, to assess barriers and facilitators to implementation over 18 months. Methods: The Consolidated Framework for Implementation Research (CFIR) was used to develop baseline and follow-up surveys. These were distributed to members of the core implementation team at the Gender Wellness Center at the midpoint and conclusion of the Robert Wood Johnson Foundation grant. Responses were largely open-ended and analyzed qualitatively. Results: Results are presented in terms of CFIR domains and constructs, as well as the relative outlook (positive or negative) of implementation. Overall, there were improvements over time, with more encouraging feedback and examples of success at follow-up. Though true, organizational culture and individual beliefs about the provision of transgender care challenged implementation of the Center of Excellence throughout the project. Conclusions: This study highlights the importance of organizational culture on implementation efforts, as well as the need for complex, multifaceted interventions to overcome such challenges in order to improve care for marginalized populations.