Embryo Freezing

Vitrification

  • Vitrification can be defined as a physical process by which a highly
  • concentrated solution of cryoprotectant solidifies into a glassy vitrified state from the liquid phase by an extreme elevation in the viscosity while cooling at a low temperature.
  • The soild which is called glass retains the normal molecular and ionic distribution of liquid state and can be considered to be an extremely viscous super cooled liquid.
  • The process avoids intracellular and extracellular ice formation.
  • Thus avoids the possible damage which can be caused by intracellular ice formation and the osmotic effects related to extracellular ice formation.
  • it also completely avoids ice crystal formation in cryopreserved cells during the thawing process.
  • Vitrification of water inside cells can be achieved by :
    • Increasing the speed of temperature conduction
    • Increasing the concentration of cryoprotectant

Vitrification-cooling

Cryoleaf Loading of the embryos on the cryoleaf
  • Take out equilibration media (vial 1) and vitrification media (vial 2) in a petridish
  • Label cryoloops with patient's ID No, Name, Date
  • Transfer the embryos to be vitrified to the equilibration media and keep alarm for 5 minutes, using the suitable flexi pipette with minimum media
  • At the end of 5 minutes,transfer embryos to Vitrification solution and load on to the cryoloop using flexi pipette
  • Hold/touch loop over the Vitrification machine till the ice formation occurs
  • Insert the loop in the cover and preserved in liquid nitrogen
  • Make the entries for location

Vitrification Warming

  • Take out patient sheet and locate the embryos
  • Take out the warming media in petridish
  • Quickly take out the loop from the cover and touch it in the warming
  • media and keep the embryos for 3 minutes
  • Transfer the embryos into dilution media 1,2,&3 respectively and keep
  • for 3 minutes in each using flexi pipette
  • Transfer embryos in pre equilibrated culture media
Tip of cryoleaf containing the embryo in warming media
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