Whole-mount in situ hybridization and Blimp1 immunohistochemistry

Whole-mount in situ hybridisation analysis was performed as before^{50 (link)}, using antisense riboprobes for Bmp2^{64 (link)}, Bmp4^{13 (link)} and Gdf3^{28 (link)}. For Blimp1 immunohistochemistry, E7.5 decidua were fixed overnight in 4% PFA, dehydrated in ethanol, embedded in paraffin wax and sectioned (6 μm). Dewaxed sections were subjected to antigen retrieval by boiling for 1 h in Tris/EDTA (pH 9.0) and permeabilized for 10 min in 0.1% Triton X-100 in TBS. Sections were subsequently blocked with 10% normal goat serum in TBS. Sections were incubated in rat monoclonal anti-Blimp1 (1:500 dilution, sc-130917, Santa Cruz Biotechnology) in block overnight at 4 °C and signal-amplified with rabbit anti-rat secondary antibody (AI-4001, Vector Laboratories) for 45 min at RT followed by peroxidase blocking for 20 min at RT and development with Envison System-HRP for rabbit antibodies (K4011, DAKO) and Vector Red substrate (SK-4805, Vector Laboratories). Sections were lightly counter stained with haematoxylin, coverslipped and imaged.
Haematoxylin and eosin staining was performed as previously described^{36 (link)}.