Background. The goal of this study was to understand prospective cohort Principal Investigators’ (PIs’) attitudes regarding the importance of religion and spirituality (R/S) on disease etiology in order to identify barriers and opportunities for greater inclusion of these domains in high-quality epidemiological research.Methods. One-hour, semi-structured qualitative interviews were conducted with 20 PIs representing 24 National Institutes of Health (NIH)-funded prospective cohort studies in the US, collectively capturing health data on 1 in 100 Americans. Sample size was calculated to achieve thematic saturation.Results. The majority of PIs we interviewed saw R/S as a potentially important area to investigate to better understand disease etiology, but some were unsure about the mechanisms through which R/S can impact health and how to translate this knowledge into public health interventions. PIs identified 5 key areas that are lacking in current R/S research that need to be addressed in future research to convince them of collecting more R/S measures in their cohorts: (1) high-quality, prospective studies; (2) examining a plausible biological mechanism; (3) commonly collected, well-validated R/S measures; (4) addressing bias against R/S research; and (5) NIH funding for R/S research.Conclusions. Results of this study provide a roadmap for future R/S research investigating the impact of R/S on disease etiology in the context of U.S. prospective cohort studies. Such research could provide potentially important information concerning how to address existing health disparities in the U.S.