The tissue sections were covered with agarose gel bond support medium (Gel Bond Bio-zyme, 0.2 mm; Edwards Instrument Commpay, Narellan, NSW, Australia) and incubated at 45°C for 16 h in a moist chamber. After hybridization, the sections were washed in double-strength SSC for 15 min; they were then washed in 300 mM NaCl, 20 mM Tris-HCl (pH 7.5), 1 mM EDTA, 10 mM dithiothreitol, and 50% formamide at 37°C for 10 min and treated with RNase A (20 mg/ml) at 37°C for 30 min. They were next washed in double-strength SSC and 0.1-strength SSC at room temperature for 10 min each time and then in 0.1-strength SSC at 45°C for 15 min. The hybridized probe was detected using a Nucleic Acid Detection kit (Boehringer Mannheim). Slides were rinsed in 10 mM Tris-HCl (pH 7.5) and 1 mM EDTA, counterstained with 0.2% methyl green, and examined under brightfield microscopy. Controls consisted of 1) hybridization with the sense riboprobe and 2) RNase A pretreatment (20 mg/ml) at 37°C for 30 min before hybridization. antibiotics levaquin
Characterization of the MTR in Male Reproductive Tract Tissues
Adult male prostate and testis tissues were weighed before homogenization in 20 (v:w) 10 mM Tris-HCl containing 1.5 mM EDTA (pH 7.4). The homogenates were centrifuged at 1000 X g for 15 min; supernatants were collected and recentrifuged at 120 000 X g for 30 min.