Quinn s advantage human htf fertilization media

All reagents to prepare non-commercial Human Tubal Fluid (HTF) and Toyoda–Yokoyama–Hosi (TYH) media, as well as valinomycin, FITC-labeled Pisum sativum agglutinin, Tween 20, 2′,7′-Bis-(2-Carboxyethyl)-5-(and-6)-Carboxyfluorescein, Acetoxymethyl Ester (BCECF-AM), and calcium ionophore A23187, were from Sigma (St. Louis, MO); 3, 3′-dipropyl- thiadicarbocyanine iodide [DiSC₃(5)] was from Invitrogen (Carlsbad, CA, United States); Hoechst 33342 was from Cayman Chemicals, (Ann Arbor, MI, United States); paraformaldehyde was from Electron Microscopy Sciences (Hatfield, PA, United States); Dulbecco’s Phosphate Buffered Saline (DPBS) was from GIBCO Life Technologies (Gaithersburg, MD, United States); and Quinn’s Advantage Human (HTF) fertilization media and human serum albumin were from CooperSurgical (Trumbull, CT, United States).

Non-commercial HEPES-buffered HTF (4.67 mM KCl, 0.31 mM KH₂PO₂, 90.69 mM NaCl, 1.20 mM MgSO₄, 2.78 mM glucose, 1.6 mM CaCl₂, 0.51 mM sodium pyruvate, 60 mM sodium lactate, 23.8 mM HEPES, and 10 μg/ml Gentamicin, (pH 7.4 with NaOH) was used to measure human sperm membrane potential. Non-commercial HEPES-buffered HTF was supplemented the same day with 25 mM NaHCO₃ and 0.5% w/v BSA A-7906 (Sigma) for capacitating conditions, and only with 0.5% w/v BSA for non-capacitating conditions. IVF samples were processed in commercial Quinn’s Advantage Human (HTF) fertilization media [CooperSurgical (Trumbull, CT, United States)] supplemented with 3 mg/ml human serum albumin (CooperSurgical) and pre-incubated overnight at 37°C with 5% CO₂ to equilibrate pH. The same media was used to capacitate IVF and donor sperm samples.
Mouse capacitating modified (TYH) medium [119.30 mM NaCl, 4.70 mM KCl, 1.71 mM CaCl₂, 1.20 mM KH₂PO4, 1.20 mM MgSO₄, 0.51 mM sodium pyruvate, 5.56 mM glucose, 20 mM HEPES, 15 mM NaHCO₃ and 5 mg/ml BSA A-2153 (pH 7.4 with NaOH)] was used to capacitate mouse sperm.