Development of Cheaper Embryo Vitrification Device Using the Minimum Volume Method

PLoS One. 2016 Feb 5;11(2):e0148661. doi: 10.1371/journal.pone.0148661. eCollection 2016.


This study was designed to compare the efficiency of the Cryotop and Calibrated plastic inoculation loop (CPIL) devices for vitrification of rabbit embryos on in vitro development and implantation rate, offspring rate at birth and embryonic and fetal losses. CPIL is a simple tool used mainly by microbiologists to retrieve an inoculum from a culture of microorganisms. In experiment 1, embryos were vitrified using a Cryotop device and a CPIL device. There were no significant differences in hatched/hatching blastocyst stage rates after 48 h of culture among the vitrified groups (62 ± 4.7% and 62 ± 4.9%, respectively); however, the rates were significantly lower (P<0.05) than those of the fresh group (95 ± 3.4%). In experiment 2, vitrified embryos were transferred using laparoscopic technique. The number of implanted embryos was estimated by laparoscopy as number of implantation sites at day 14 of gestation. At birth, total offspring were recorded. Embryonic and fetal losses were calculated as the difference between implanted embryos and embryos transferred and total born at birth and implanted embryos, respectively. The rate of implantation and development to term was similar between both vitrification devices (56 ± 7.2% and 50 ± 6.8% for implantation rate and 40 ± 7.1% and 35 ± 6.5% for offspring rate at birth); but significantly lower than in the fresh group (78 ± 6.6% for implantation rate and 70 ± 7.2% for offspring rate at birth, P<0.05). Likewise, embryonic losses were similar between both vitrification devices (44 ± 7.2% and 50 ± 6.8%), but significantly higher than in the fresh group (23 ± 6.6%, P < 0.05). However, fetal losses were similar between groups (10 ± 4.4%, 15 ± 4.8% and 8 ± 4.2%, for vitrified, Cryotop or CPIL and fresh, respectively). These results indicate that the CPIL device is as effective as the Cryotop device for vitrification of rabbit embryos, but at a cost of €0.05 per device.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cryopreservation / economics
  • Cryopreservation / methods
  • Cryopreservation / veterinary*
  • Embryo Culture Techniques
  • Embryonic Development*
  • Female
  • Pregnancy
  • Pregnancy Outcome / veterinary
  • Rabbits
  • Vitrification

Grants and funding

This research was supported by the projects Spanish Research project AGL2014-53405-C2-1-P Comisión Interministerial de Ciencia y Tecnologia (FMJ, JSV) and Generalitat Valenciana research program (Prometeo II 2014/036, JSV, FMJ).