Sensitive methods to detect epithelial disruption: tests for microhemorrhage in cervicovaginal lavages

The toxic effects of topical microbicides might include epithelial disruption, and if sufficiently severe, may cause visible bleeding. We have developed two sensitive methods to detect hemorrhage far below the visual detection threshold: an enzyme-linked immunosorbent assay for human hemoglobin and a method for staining and enumerating red blood cells by fluorescent microscopy. We have applied these methods to cervicovaginal and introital lavages, and found them to be sensitive and quantitative over a broad range. Comparing the new techniques with a standard heme-detection assay (Hemastix) we found them to be more sensitive, more objective, more fully quantitative, and more likely to be indicative of recent events. In pilot studies we found that the red blood cell and hemoglobin concentrations in samples taken in the luteal phase of the menstrual cycle are low, but still within the detection range of the new assays, providing a quantifiable baseline from which increases in blood from epithelial disruption can be detected and quantified. We propose that these new methods may be used for the safety assessment of microbicides by detecting and quantifying microhemorrhage in cervicovaginal secretions before and after exposure to microbicide.