Ika ultra turrax homogenizer

Levels of E2 and P4 in luteal tissue in response to intrauterine treatment with E2 were determined by radioimmunoassay as described previously [15 (link),52 (link)]. Briefly, luteal tissues were homogenized in 5% trichloroacetic acid (1 mL of 5% TCA per 1 mg of luteal tissue) using IKA Ultra-Turrax homogenizer (IKA^®-Werke GmbH & Co. KG; Staufen, Germany). Homogenates were then centrifuged (10,000× g, 10 min, 4 °C) and supernatants were collected and neutralized with NaOH (2.5 mM) to pH 7.5. The abundance of E2 and P4 was then determined using radioimmunoassay kits (DIAsource, Louvain-la-Neuve, Belgium) accordingly to the manufacturer’s protocol. The sensitivity of the E2 and P4 assays were 3 pg/mL, and 0.05 ng/mL, respectively. The intra-assay coefficient of variations (CV) for the E2 and P4 assays were 6.8% and 5.5%, respectively.

Animals were anesthetized using 3% pentobarbital sodium 30 mg/kg injected via the marginal ear vein. The cortex and hippocampus of the rabbits were separated after the brains were extracted. A 10% w/v rabbit cortex and hippocampus homogenate were made in pre-chilled physiological saline using an IKA ULTRA-TURRAX homogenizer (IKA®-werke Gmbh & Co., KG, German), the obtained homogenate was centrifuged at 4500 rpm, 4 °C for 10 min. The supernatant was extract and the Aβ1–42 level was assayed immediately using homologous specific ELISA kits (Nanjing Jiancheng Bioengineering Institute, China), according to the manufacturer’s instructions. Protein concentration was determined by Coomassie brilliant blue method (Nanjing Jiancheng Bioengineering Institute, China).