R18S3 (Medium for Mouse Sperm Cryopreservation)

      4. Put the sealed straw into float, with turning sperm-suspension layer to the bottom.Put the straw-fl oat on the surface of liquid nitrogen in N2 tank.  
          Keep it stand for 15 min.

      5. After 15 min, soak it into liquid nitrogen.

Thawing


1. Pick up the freezing straw from liquid nitrogen, and soak it into warm water at 37 ℃ .
    Keep it stand for 15 min.

2. After 15 min, pick up the straw from warm water and wipe it with some papers.
    After that, cut the both edges of straw and drop the sperm-suspension on the petri dish.

3. Pick up 2 µL of sperm-suspension and apply it into 200 µL HTF, which is pre-warmed in incubator
    (37 ℃ , 5% CO2 in air) beforehand.
    After 1 hour from above treatment, observe the motility and active level.