In this work, we used 0.6% acetic acid, in vivo 3% to 5% acetic acid is typically applied to the cervix. The epithelium of the cervix, like all epithelia, is a barrier that regulates the flow of molecules. The ectocervix is comprised of stratified squamous epithelium. The top layer of the epithelium is comprised of glycogen containing, cornified cells.27 Some cornified epithelial layers provide significant protection to the underlying epithelial cells,28 however, we have not been able to find proof of this for the specific case of the cervical epithelium. A few cell layers down tight junctions between cells restrict the movement of molecules passing between the cells.27 The concentration of acetic acid is likely much less in the lower layers of the epithelium. This view is supported by modeling of acetic acid in the epithelium.29 While we do not know the exact concentration of acetic acid in the epithelium, we do know that fairly low concentrations of acetic acid can cause permanent damage to cells in vitro. We performed measurements of the ability of MR1 and SiHa cells to continue growing in vitro after application of acetic acid which demonstrated that after only 5 min in 0.3% acetic acid, the ability of these cells to grow was greatly reduced (unpublished data). Similarly, Wu and Qu12 used 0.3% and 0.6% acetic acid in their work and reported that permanent cell damage occurred when concentrations of 1.2% or more was used. Therefore, we chose a concentration of acetic acid that was expected to provide a large affect, but not immediately kill the cells. When comparing the concentration of acetic acid used in this work to that used in vivo, another important factor is that the buffering capicity of cellular tissue is likely very different from that of cell culture. There are many more cells in a given volume and fluid movement in the epithelium and blood flow under the epithelium may also provide additional pH buffering. A low concentration of acetic acid in vitro may have effects on cells similar to those that occur when a higher concentration of acetic acid is used in vivo.