A description of the polyclonal rabbit anti-rat MCP-1 antibody used for immunohistochemical staining has been published. The monoclonal antibody against rat monocytes/macrophages (clone ED1) used for immunohisto-chemical staining was obtained from Chemicon International Inc. (Temecula, CA). The specificity of this antibody for rat monocytes/macrophages has been reported previously. The monoclonal antibody against rat differentiated macrophages (clone ED2) was obtained from Serotec via Accurate Chemical & Scientific Corp. (Westbury, NY) and has been shown to specifically recognize differentiated, resident tissue macrophages. Biotinylated secondary antibodies to mouse and rabbit immunoglobulin were obtained from Vector Laboratories (Burlingame, CA). ampicillin antibiotic
Frozen sections of ovaries or isolated corpora lutea were air-dried and fixed in 95% ethanol (10 min); they were then placed in 0.3% H2O2 in methanol (4°C) for 15 min (macrophages) or 30 min (MCP-1) to quench endogenous peroxidase activity. The tissue sections were rinsed three times (5 min each) in PBS containing 1% BSA (fraction V; PBS-1% BSA) and then incubated with 10% normal goat serum for 15 min (macrophages) or 30 min (MCP-1) at 37°C.