BACKGROUND: Sexually transmitted infection (STI) surveillance is vital for tracking the scale and pattern of epidemics; however, it often lacks data on the underlying drivers of STIs. OBJECTIVE: This study aimed to assess the acceptability and feasibility of implementing a bio-behavioral enhanced surveillance tool, comprising a self-administered Web-based survey among sexual health clinic attendees, as well as linking this to their electronic health records (EHR) held in England's national STI surveillance system. METHODS: Staff from 19 purposively selected sexual health clinics across England and men who have sex with men and black Caribbeans, because of high STI burden among these groups, were interviewed to assess the acceptability of the proposed bio-behavioral enhanced surveillance tool. Subsequently, sexual health clinic staff invited all attendees to complete a Web-based survey on drivers of STI risk using a study tablet or participants' own digital device. They recorded the number of attendees invited and participants' clinic numbers, which were used to link survey data to the EHR. Participants' online consent was obtained, separately for survey participation and linkage. In postimplementation phase, sexual health clinic staff were reinterviewed to assess the feasibility of implementing the bio-behavioral enhanced surveillance tool. Acceptability and feasibility of implementing the bio-behavioral enhanced surveillance tool were assessed by analyzing these qualitative and quantitative data. RESULTS: Prior to implementation of the bio-behavioral enhanced surveillance tool, sexual health clinic staff and attendees emphasized the importance of free internet/Wi-Fi access, confidentiality, and anonymity for increasing the acceptability of the bio-behavioral enhanced surveillance tool among attendees. Implementation of the bio-behavioral enhanced surveillance tool across sexual health clinics varied considerably and was influenced by sexual health clinics' culture of prioritization of research and innovation and availability of resources for implementing the surveys. Of the 7367 attendees invited, 85.28% (6283) agreed to participate. Of these, 72.97% (4585/6283) consented to participate in the survey, and 70.62% (4437/6283) were eligible and completed it. Of these, 91.19% (4046/4437) consented to EHR linkage, which did not differ by age or gender but was higher among gay/bisexual men than heterosexual men (95.50%, 722/756 vs 88.31%, 1073/1215; P<.003) and lower among black Caribbeans than white participants (87.25%, 568/651 vs 93.89%, 2181/2323; P<.002). Linkage was achieved for 88.88% (3596/4046) of consenting participants. CONCLUSIONS: Implementing a bio-behavioral enhanced surveillance tool in sexual health clinics was feasible and acceptable to staff and groups at STI risk; however, ensuring participants' confidentiality and anonymity and availability of resources is vital. Bio-behavioral enhanced surveillance tools could enable timely collection of detailed behavioral data for effective commissioning of sexual health services.