Immunohistochemistry of Human Ovarian DDC

Sections of human ovaries derived from a local collection at Anatomy III, Cell Biology (Munich, Germany) were used for immunohistochemistry. Immunohistochemistry was performed with a rabbit antiserum raised against human DDC (Sigma-Aldrich) and the corresponding blocking peptide (Sigma-Aldrich). Tissue samples and immunohistochemistry were described previously [5 (link), 18 (link)]. In brief, after removal of paraffin, antigen retrieval and blocking of endogenous peroxidase activity, the tissue was incubated in 5 % appropriate serum, diluted in phosphate-buffered saline. Antiserum incubation was done overnight at 4 °C. The antiserum against human DDC was diluted 1:50. Afterwards, incubation with a biotinylated secondary antibody (1:500 dilution; Dianova, Hamburg, Germany) for 2 h at room temperature was performed. A Vectastain ABC Kit (Vector Laboratories, Burlingame, CA, USA) and a 3,3′-diaminobenzidine tablet set (Sigma-Aldrich) were used for the final staining procedure. Slides were covered with Entellan (Merck Millipore, Billerica, MA, USA). For control purposes, incubation with antigen peptide pre-adsorbed antiserum was employed.