Dakocytomation liquid dab substrate chromogen system

Prostate ventral lobes and the bladder were harvested for histological analyses after cystometry. One part of the prostate and the bladder was fixed in buffered 10% formaldehyde solution for 24 hr, embedded in paraffin, cut with a microtome, and stained with hematoxylin-eosin for evaluating tissue inflammation. The remainder of the prostate was used for immunohistochemical staining. Paraffin embedded prostate sections were placed on silicone-coated slides. After deparaffinization in xylene and rehydration using graded alcohol solutions, the sections were fixed with 10 mM sodium citrate pH 6 at 105°C by Autoclave (TOMY SEIKO co. Ltd.) for antigen retrieval. The tissues were rinsed with phosphate buffered saline (PBS), transferred to 0.3% hydrogen peroxide for 10 min to block peroxidase activity, and rinsed with distilled water and PBS. The sections were blocked with 10% normal goat serum (NICHIREI CORPORATION.) for 30 min. These tissues were incubated with an anti-ERβ antibody (1:1,000; Santa Cruz Biotechnology, Inc.) for 1 hr at room temperature. After washing with PBS, the tissue sections were incubated for 30 min with HRP-labeled polymer anti-rabbit antibody (DAKO.) at room temperature. After washing with PBS, the color was developed using the Dako Cytomation Liquid DAB Substrate Chromogen System (DAKO). The tissues were counterstained with hematoxylin.