The human cervicovaginal mucosa is the primary target of HIV-1 infection during male to female transmission. This tissue contains the the full spectrum of cell types and immune modulators that comprise both the innate and adaptive arms of the immune system. Mounting evidence indicates that mucosal epithelial cells are sentinels of the female reproductive tract, producing innate immune mediators that control the vaginal microflora under normal conditions. Recent studies, however, indicate that certain factors secreted in response to another pathogen or after exposure to a vaginal product may in fact enhance infection by HIV-1. Mucosal inflammation and CD4 cell activation as well as disruption of TLR function and epithelial integrity represent potential causes for such effect. It is therefore important to make sure that vaginal products, including microbicides, do not disrupt the structure or function of the cervicovaginal mucosa. Although a number of biomarkers have been linked to microbicide-induced cervicovaginal inflammation and many of these markers have been measured in preclinical and clinical assays, there are currently no data that demonstrate a correlation between any one marker and susceptibility to HIV-1 infection in humans. To date, the lack of a validated biomarker of cervicovaginal safety represents a gap in the knowledge base that hinders the rational and expeditious selection of microbicide candidates entering clinical trials. Current discovery efforts and preclinical assessment of microbicide safety use an integrated sequential evaluation system that includes cell-based models, explant-based models, and animal-based models. Relevant research in these areas is yielding new assays and biomarkers that, if validated, will be essential to the rational selection of microbicide candidates for efficacy trials.