To Freeze Giardia in cryovials
- Prepare a separate solution of sterile filtered 18% DMSO in culture medium (e.g., take 4.1 ml medium, add 0.9 ml DMSO and sterile filter using a syringe filter).
- Label liquid nitrogen vials with name of cell line, strain of Giardia, date and your initials. Record this in culture book.
- Ice cells for 15 minutes.
- Spin down 12mL of confluent culture for 5 min at (900 x g) @4 degrees Celsius.
- Pour off all supernatant and add 1ml of fresh culture medium.
- Add 1ml 18% DMSO to 1ml resuspended culture for a final [9%]. Mix gently and aliquot into two liquid nitrogen vials using a sterile transfer pipet
- Transfer to –80°C for several days. Move to liquid nitrogen tank for permanent storage.
To thaw Giardia from cryovial
- Bring stored vial rapidly to 37°C. Record removal of frozen aliquot in the culture book.
- Pour into a 15ml culture tube with fresh medium, and let attach at 37°C for about 60 minutes.
- Pour off supernatant and replace with fresh medium (no DMSO). Add puromycin at this point, if necessary.
- Culture will be confluent the following day.