Evaluation of a novel microfluidic chip-like device for purifying bovine frozen-thawed semen for in vitro fertilization

The aim of this study was to validate a novel sperm purification device, the VetCountTM Harvester, for use in bovine in vitro fertilization (IVF). The device's performance was compared to BoviPureTM gradient centrifugation, a commercially available and accepted routine technique. Semen quality parameters were assessed for frozen-thawed semen from six different bulls (n = 6) following sperm purification. For each bull two semen subsamples were purified utilizing BoviPureTM gradient centrifugation and the Vet-CountTM Harvester, including a third subsample as untreated control. Both treatments significantly increased the proportion of progressively motile sperm cells (84.4 & PLUSMN; 14.1% and 85.1 & PLUSMN; 7.8%, respectively) compared to the untreated semen (41.9 & PLUSMN; 18.8%). BoviPureTM gradient and VetCountTM Harvester selected predominantly viable acrosome intact (VAI) sperm cells with low membrane fluidity and low free intracellular calcium concentration [Ca2+]i (76.5 & PLUSMN; 4.4% and 78.6 & PLUSMN; 6.0%). Normalizing [Ca2+]i of VAI sperm cells (non-treated semen: [Ca2+]i = 1) VetCountTM Harvester purified spermatozoa (0.67 & PLUSMN; 0.10) showed significantly lower [Ca2+]i than BoviPureTM treated sperm (0.84 & PLUSMN; 0.14; P < 0.05). Subsequently, the fertilizing ability of the spermatozoa was evaluated performing a competitive fertilization assay. Sperm cells from both treatment groups were fluorescently labelled using different dyes and added in equal amounts to in vitro matured oocytes. After 18 h co-incubation, the origin of the fertilizing sperm cell was evaluated via fluorescence microscopy. In two bulls, VetCountTM Harvester selected sperm that fertilized significantly more oocytes then BoviPureTM treated sperm, in another bull it was the opposite. For three bulls no difference was observed.We conclude that the VetCountTM Harvester selects a high-quality, fertile sperm fraction from frozen-thawed bull semen. However, some considerations have to be kept in mind for the direct use of the isolated sperm fraction in IVF.

Automated summary

The VetCountTM Harvester was compared to the BoviPureTM gradient centrifugation for purifying sperm in bovine IVF. Both methods significantly increased the proportion of motile sperm compared to untreated semen. The VetCountTM Harvester selected viable acrosome intact sperm cells with lower [Ca2+]i compared to BoviPureTM treated sperm. The fertilizing ability of the spermatozoa varied, with VetCountTM Harvester showing higher fertility in some cases.