Optimization of the in vitro fertilization protocol for frozen epididymal sperm with low fertilization ability in Ban—A native Vietnamese pigs

Abstract

The aim of the present study was to improve the penetration during in vitro fertilization (IVF) of a frozen lot of epididymal sperm with a notoriously low fertilization ability of a Ban boar which is a native Vietnamese breed by optimizing different parameters of the IVF system. In Experiment 1, we determined that Pig-fertilization medium was superior medium to Tyrode's albumin lactate pyruvate-polyvinyl alcohol medium for IVF and defined the optimum the sperm concentration (1 × 10 sperm/ml). In Experiment 2, we clarified that partial removal of cumulus cells from cumulus-oocyte complexes by hyaluronidase treatment before IVF enhances sperm penetration, whereas complete cumulus removal reduces penetration. Finally, in Experiment 3 the elevation of concentration of caffeine in Pig-fertilization medium from 2 to 5 mmol/L and the prolongation of the co-culture of gametes from 3 to 5 hr significantly increased the total penetration rate from 15.2% to over 50%. In conclusion, the combination of partial oocyte denudation, an elevated caffeine concentration in Pig-fertilization medium and an extended interval of IVF with using an optimized sperm concentration was a potent way to improve the fertilization results for a frozen epididymal Ban sperm lot with low fertility.

Optimization of in vitro embryo production and zygote vitrification for the indigenous Vietnamese Ban pig: The effects of different in vitro oocyte maturation systems

“…study we employed an IVF protocol which has been optimized recently for the efficient utilization of frozen epididymal Ban sperm (Linh et al, 2018).…”

“…The reason for these different results might be the different laboratory conditions and protocols between the studies. In the current study we employed an IVF protocol which has been optimized recently for the efficient utilization of frozen epididymal Ban sperm (Linh et al., 2018).…”

“…The IVF procedure was performed according to the method of Linh et al. (2018). The medium used for IVF was Pig‐FM containing 90 mM/L NaCl, 12 mM/L KCl, 25 mM/L NaHCO 3 , 0.5 mM/L, NaH 2 PO 4 , 0.5 mM/L MgSO 4 , 10 mM/L sodium lactate (Kanto Chemical Co., Inc., Tokyo, Japan), 10 mM/L HEPES, 8 mM/L CaCl 2 , 2 mM/L sodium pyruvate (Sigma), 2 mM/L caffeine, and 5 mg/ml Bovine Serum Albumin (BSA, fraction V; Sigma).…”

Production of Agu piglets after transfer of embryos produced in vitro

“…The in vitro fertilization (IVF) procedure was performed according to the reports of Kikuchi et al (2002) and Linh et al (2018) with slight modifications. After IVM, all COCs were treated for a short time with 0.1% hyaluronidase (Sigma‐Aldrich) in collection medium to remove outer layers of cumulus cells.…”

Cryopreservation of immature oocytes of the indigeneous Vietnamese Ban Pig

“…The IVF procedure was performed according to the report of Linh et al (2018). The medium used for IVF was a modified Pig-FM (Kikuchi et al, 2002;Suzuki et al, 2002) medium containing 10 mM HEPES, 5 mM caffeine, and 5 mg/ml BSA.…”