Veterinarian View

contributed article by Dr. Vince Collison, Collison Embryo | Collison Veterinary Services, Rockwell City, IA

One thing that makes me cringe is seeing an owner pulling canes way out of the neck of a nitrogen tank to look to see what it is prior to transfer or AI. The first key to good results with frozen embryos or semen is to keep them frozen until the time you are ready to thaw them.

When an embryo is cryopreserved, it has to remain in the frozen state at all times to maintain integrity. Some of the numbers to remember for handling frozen embryos and semen are these:

This is the temperature of liquid nitrogen. This is the temperature where all metabolism stops on cells and any intracellular ice crystals are stable. This is also the temperature of the body of a liquid nitrogen tank where all the goblets are stored. A tank can get to a fairly low level of liquid nitrogen and still maintain this temperature in the body of the tank.

This is the highest temperature where ice crystals within the cryopreserved cells stay stable. If temperatures go above this level on frozen embryos or semen, there will be recrystallization of ice crystals inside of the cells. This recrystallization process causes shearing of cell membranes resulting in irreversible damage. The straws in this case can be warmed up above this level and damage occurs without ever being totally thawed.

This is the temperature in the neck of the nitrogen tank.

The other thing to be aware of is a ¼ cc straw is more prone to thaw damage than a ½ cc straw since the surface area relative to the volume is higher in the ¼ cc straw. As semen becomes more commonly packaged in ¼ cc straws it will be more important to handle semen correctly for best results. Also all embryos are packaged in ¼ cc straws, so they are at higher risk for thawing.

For people that store their own embryos we recommend these guidelines:

1) Keep the nitrogen tanks at least 50% full at all times. This will keep the lower goblet submerged in nitrogen and also help reduce the risk if you pull the canister up into the neck of the tank.

2) Keep good records of cane codes so you know what each cane contains.

3) Use a flashlight to look down into the tank when finding embryos rather than pulling the canister way up into the neck of the tank.

4) If you have to split embryos between canes, do this with the cane submerged in liquid nitrogen rather than in the neck of the tank exposed to the air temperature. We pour liquid nitrogen into upright Igloo coolers to do this. It allows you to read the labels easily and transfer straws between canes while submerged in nitrogen.

5) When transferring canes from a vapor shipper to a regular nitrogen tank, pour some nitrogen into the vapor shipper before making the transfer between tanks. This fills the goblet with some liquid nitrogen and gives the embryo some protection from thawing when moving to a regular tank. Vapor shippers are designed to keep embryos and semen cold, but there is no free liquid in the tank. So if you don’t pour some liquid nitrogen in the shipper to fill the cane goblet, the straws will be exposed to air during the move between tanks. Once this is done, pour the liquid back out of the vapor shipper. For the shipper to be sent back to the original destination, it cannot have any free liquid nitrogen in it.

Proper handling of frozen semen and embryos can make all the difference between good and bad results. Stick to these basic guidelines to ensure best results.

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