An effective vaginal microbicide formulation must distribute and maintain an epithelial coating layer. The post-application durability of this coating is significantly affected by the vaginal environment. A new in vitro assay quantified coating layer erosion after contact and shear with simulated vaginal fluid or semen. Coating layer persistence and viscosity of both fluid and gel layers were assessed versus time. Five vaginal formulations were studied. In all gels, there was an overall trend of rapid ( approximately 30 min) and significant viscosity loss. Although there were differences across gels and between simulants, greater erosion occurred after contact with the low-pH vaginal fluid simulant (>50% viscosity decrease), as compared to an alkaline semen simulant. These in vitro results suggest significant differences in vivo of vaginal coating retention by the test gels. This new assay can be diversified to create a spectrum of biologically relevant conditions which collectively simulate the natural history of vaginal formulation residence.