Benzylpenicillin potassium

All reagents and solvents purchased were the highest commercial quality and used without further purification otherwise noted. Dimethyl sulfoxide (DMSO) was purchased from KANTO chemicals. Propidium iodide (PI), benzylpenicillin potassium, and streptomycin sulfate were purchased from Fujifilm Wako Chemicals. 1,3-Diphenylisobenzofuran (DPBF) was purchased from Sigma-Aldrich. Minimum essential medium (MEM) and phosphate-buffered saline (PBS) were purchased from Nacalai Tesque. Fetal bovine serum (FBS) was purchased from Capricorn Products Inc. The Ir complex 7 was synthesized according to our previous paper [26 (link)], a stock solution of which in DMSO was prepared and stored at 0 °C prior to use. UV-Vis absorption spectra of DPBF were measured on a JASCO V-550 UV-vis spectrophotometer. The microscopic images of HeLa S3 cells were observed on fluorescent microscopy (Biorevo, BZ-x800, Keyence, Amagasaki, Japan).

Porcine ovaries were obtained from a local slaughterhouse and transported to the laboratory in 0.9% saline containing 0.75 µg/mL benzyl-penicillin potassium (Wako, Osaka, Japan) and 0.5 µg/mL streptomycin sulfate salt at 37–38 °C. After the ovaries had been washed, cumulus oocyte complexes (COCs) were aspirated from follicles (diameter: 3–8 mm) using a 10 mL syringe with an 18-gauge needle. COCs with three or more layers of cumulus cells and homogeneous cytoplasm were selected and washed three times in 0.9% saline with 1 mg/mL bovine serum albumin (BSA). Washed COCs were incubated in IVM I medium for 22 h at 38.5 °C and 5% CO₂ in air. During the first period of maturation (0–22 h), the IVM I medium consisted of 10% porcine follicular fluid, 0.57 mM cysteine, 25 μM β-mercaptoethanol, 10 ng/mL epidermal growth factor, 10 IU/mL pregnant mare serum gonadotropin (Prospec, East Brunswick, NJ, USA), and 10 IU/mL human chorionic gonadotropin (Prospec). After the first maturation period, a second period (from 22 to 44 h) was initiated. The same media was used (without hormone).