Background This is the first study, of which we are aware, to evaluate the feasibility and accessibility of simultaneous use of Global Positioning System (GPS) and saliva collection for biomarker assessment as an objective measure of stress physiology among sexual minority (lesbian, gay, bisexual, queer, and other non-heterosexual identities) individuals. The principal motivation for pairing GPS and saliva collection was to investigate how characteristics of the built and social environments along with participants’ daily activity paths affect stress. This can contribute to a better understanding of health and health behaviors in the sexual minority community. Methods A convenience sample of enrolled participants (N = 124) from Houston, Texas was asked to complete questionnaires, carry with them a GPS unit daily, and collect and store 6 samples of saliva at specific times across the span of a day prior to a second visit around one week later. Results Of 124 participants, 16 participants (12.90%) provided no useable GPS data and 98 (79.03%) provided at least 4 days of data. More than three-fourths (n = 98, 79.03%) also provided complete saliva samples. Conclusions Our results show that the simultaneous use of GPS and saliva collection to assess sexual minority individuals’ activity paths and stress level is feasible.