In situ zymography
Areas with gelatinase activity were detected by in situ zymography as previously described (Khandoker et al. 2001; Nemori and Tachikawa 1999). GN film (Fuji Photo Film Co.) coated with gelatin base emulsion was used to detect the site of gelatinase activity in the underlying tissue. Unfixed 7-μm-thick ovarian cryosections were mounted onto the GN film, incubated for 24 h at 37 °C, and stained with Bearish Scarlet (Chroma-Gesellschaft, mbH & Co., Munster, Germany) and hematoxylin. Specific pattern of gelatin digestion was indicated by white colour due to weaker staining with BS.