The study aimed to document factors affecting willingness to engage in biospecimen donation for substance use research among Black and Hispanic/Latinx sexual and gender minority (SGM) people. From May to July 2022, we interviewed 22 Black and Hispanic/Latinx SGM people from The PRIDE Study, a cohort of SGM people in the United States and its territories, using Zoom videoconferencing software. Fourteen participants were sexual minority people, and eight participants were gender minority people. We took an inductive, collaborative qualitative analytic approach to identify themes. Themes included: (1) community benefits, (2) personal benefits, (3) community exploitation, (4) personal risks, (5) convenience, (6) trustworthiness of the research team, (7) perceived value in donating, and (8) normalization of biospecimen collection. Participants were generally motivated to engage in biospecimen donation for altruistic purposes. The most cited concerns were related to data security, misuse, and privacy. Researchers must be proactive in building trust with Black and Hispanic/Latinx SGM communities to increase engagement, diversify biospecimen repositories and reduce health inequities. Future research involving biospecimens should provide biospecimen education during the consent process and prioritize participant convenience.