Vaginal cytology for estrous cycle assessment.

Vaginal cytology was used to monitor estrous cycle stage in the female rats over a 14-day period to determine whether diet altered cycling patterns. Female Wistar rats were wrapped in a bench pad, with the animal’s vaginal opening exposed. A 2-ml disposable pipette was used to flush the vaginal cavity with ~200 μl of phosphate-buffered saline (PBS). The resulting sample was smeared on a slide and then immediately imaged (while still wet) under a light microscope at ×10 magnification. Cycle stage was determined by the presence and quantity of round cells, needle cells, and/or neutrophils (6). Male rats were wrapped in a bench pad on the same days the female rats were subjected to lavage.