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Introduction: Self-collected samples (SCS) for sexually transmitted infection (STI) testing have been shown to be feasible and acceptable in high-resource settings. However, few studies have assessed the acceptability of SCS for STI testing in a general population in low-resource settings. This study explored the acceptability of SCS among adults in south-central Uganda. Methods: Nested within the Rakai Community Cohort Study, we conducted semi-structured interviews with 36 symptomatic and asymptomatic adults who self-collected samples for STI testing. We analyzed the data using an adapted version of the Framework Method. Results: Overall, participants did not find SCS physically uncomfortable. Reported acceptability did not meaningfully differ by gender or symptom status. Perceived advantages to SCS included increased privacy and confidentiality, gentleness, and efficiency. Disadvantages included the lack of provider involvement, fear of self-harm and the perception that SCS was unhygienic. Most participants preferred provider-collected samples to SCS. Nevertheless, almost all said they would recommend SCS and would do it again in the future. Conclusion: Despite a preference for provider-collection, SCS are acceptable among adults in this setting and support expanded access to STI diagnostic services.