Embryomax ksom aa with d glucose and phenol red

The activation medium was calcium- (Ca²⁺-) free HTF complemented with 10 mM strontium chloride (SrCl₂) and 5 μg/mL cytochalasin B [40 (link)]. After being thoroughly washed in activation medium for three times, MII oocytes were activated in activation medium for 2.5 hours and then in regular HTF without SrCl₂ for 3.5 hours at 37°C with 5% CO₂. Oocytes were then transferred from activation medium to KSOM plus (+) amino acids (KSOM/AA) medium (EmbryoMax® KSOM + AA with D-glucose and phenol red, EMD Millipore, Billerica, MA, USA). Development of embryos to the 2-cell, 4-cell, morula, and blastocyst stages was assessed 24, 48, 96, and 120 hours, respectively, after the start of initial culture in KSOM/AA medium.

Mice were injected with 10 IU PMSG, followed by 10 IU human chorionic gonadotrophin (hCG, Ningbo Sansheng Pharmaceutical, Ningbo, China) 48 h later to collect MII oocytes. At 13–14 h post-hCG injection, MII oocytes were collected from the oviduct ampulla and collected in M2 medium, the cumulus cells were removed using 0.1% (w/v) hyaluronidase. Meanwhile, Different concentrations of SAL (0, 10, 20, 40 μM) were added into the medium and treated at different times (6, 12, 18, 24 h). For parthenogenesis activation, denuded oocytes were transferred first into (Ca²⁺)-free human tubal fluid (HTF) medium contained with 10 mmol/L strontium chloride and 5 μg/mL cytochalasin B (Merck, Darmstadt, Germany), cultured at 37 °C with 5% CO₂ for 2.5 h. Then oocytes were transferred into HTF with 5 μg/mL cytochalasin B, incubated at 37 °C with 5% CO₂ for 3.5 h. Activated oocytes were then cultured in KSOM plus (+) amino acids (KSOM/AA) medium (EmbryoMax^® KSOM + AA with D-glucose and phenol red, EMD Millipore, Billerica, MA, USA) at 37 °C with 5% CO₂ for early embryo development. Different stages of embryonic development (2-cell, 4-cell, morula and blastocyst) were recorded at 24, 48, 96, and 120 h, respectively.