In this video, we describe the procedure for vitrification of harvested rabbit embryos at desired development stage using ethylene glycol and dimethyl sulfoxide as cryoprotectants and cryogenic straw as the carrier system. The vitrified embryos have high survival rate and can be stored for an extended period until use.
Protocol
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Embryo Vitrification Perform all the manipulations at room temperature (around 22 °C) to reduce the vitrification solution toxicity at warmer temperatures. NOTE: Embryos can be moved using 0.1–2 µL automatic pipette in this protocol, but other similar devices to move the embryos d…
Representative Results
Figure 1: Schematization of correctly loaded straw. A) BM refers to the embryo manipulating media employed during vitrification. Embryos must be loaded in vitrification solution. B) Macroscopic appearance of the loaded straw with a magnified detail of the embryo position. This large-volume device allows us to vitrify large number of embryos, unlike minimum volume devices. Further…
Divulgations
The authors have nothing to disclose.
Materials
Bovine Serum Albumin (BSA)
VWR
332
Dimethyl Sulfoxide
Sigma Aldrich
W387509
Dulbecco's phosphate-buffered saline (DPBS)
Sigma Aldrich
D5773
Without calcium chloride.
Ethylene Glycol
Sigma Aldrich
102466-M
Liquid Nitrogen
Air Liquide
P1505XXX
Plastic Straw 0.25 mL
IMV – technologies
6431
Stereomicroscope
Leica
MZ16F
There are cheaper options such as Leica MZ8 or Nikon SMZ-10 or SMZ-2B, to name a few.
Rabbit Embryo Vitrification Using Cryogenic Straw: An Ultra-Rapid Cooling Technique to Preserve Rabbit Embryos for Long-term Storage. J. Vis. Exp. (Pending Publication), e20935, doi: (2023).