Fig 1.
Diagram showing the performance and weaknesses of the device, completed devices, and blastocysts.
Fig 2.
Vitrification devices with hollow fiber tube.
(a) Mouse 2-cell embryos were aspirated into the HFT. (b) Pipette tip attached with syringe. (c) An HFT with embryos was inserted into a pipette tip. Arrow indicates the HFT with embryos. (d) The syringe was immersed into LN2. (e) Immediately after removal from LN2. (f) The HFT inside the pipette tip was inserted into the port of an OptiCell. (g) Solution/medium exchange using a 10-ml syringe via the port. (h) OptiCell (left) and Mini-plate (right). (i) High magnification of the Mini-Plate. (j) Solution/medium exchange via the port. (k) Cell Experimental Unit (CEU). (l) Disassembled CEU. (m) New Cell Experimental Unit (NCEU). (n) The NCEU was equipped with a silicone flipper (arrow) used to pinch the HFT (arrowhead) onto the center of the plate. (o) Solution/medium exchange. (p) Blastocysts inside the HFT. Embryos were harvested from the NCEU after 4 days.
Table 1.
Recovery and blastocyst rates of embryos frozen and thawed in new devices using a hollow fiber tube.
Fig 3.
High osmolarity vitrification using a Frozebag and mesh sheet.
(a) Mesh sheet and embryos (arrows). (b) Mesh bag. (c) Frozebag before modification. (d) Equilibration with CZB medium in an incubator (5% CO, 37°C). (e) Mesh bag on the scoop. (f) Incubation of the Frozebag (5% CO2, 37°C). (g) Solution/medium exchange using 30 ml and 50-ml syringes.
Table 2.
Recovery and blastocyst rates of embryos frozen and thawed with frozebag and mesh bag.
Fig 4.
Embryo Thawing and Culturing unit (ETC).
(a) Frozebag with mesh cap. (b) Frozebag with mesh wall. (c) Blastocyst collected from the Frozebag. Although the quality of this embryos was insufficient, this was the first time that an embryo was thawed, washed, and cultured for 4 days without direct contact. (d) The top of cryotube was cut to produce the smallest tube (V-tube). (e) Protocol for thawing, washing, and culturing embryos in the ETC. (f) Blastocysts were harvested from the ETC after culture for 4 days. (g) Healthy offspring developed from the morulae/blastocysts derived from 3 days cultured embryos in ETC.
Table 3.
Recovery and blastocyst rates of embryos frozen and thawed in ETC using a cryotube or V-tube.
Table 4.
Production of offspring from embryos thawed and cultured in the ETC.
Table 5.
Development of blastocysts obtained from embryos cryopreserved at −80°C for up to three months and performed by inexperienced people.