Improving the post-thaw survival of mammalian blastocyt or sperm by applying hydrostatic pressure prior to cryopreservation
The present invention relates to a method for improving post-thaw survival of cryopreserved biological material comprising applying hydrostatic pressure to said biological material; keeping the said biological material at the hydrostatic pressure for a predetermined time period; releasing the hydrostatic pressure; and freezing the said biological material using any protocol applicable thereto. The invention also relates to the use of a pressurizing device for the pretreatment of a biological material that is to be cryopreserved, as well as to a pressurizing device for the pretreatment of a biological material that is to be cryopreserved, said device comprising a pressure chamber for receiving biological material, means to produce said pressure, and means to maintain said pressure in said chamber.
1. A method for improving post-thaw survival of cryopreserved viable biological material comprising:
(a) applying hydrostatic pressure in the range of 10 to 200 MPa to said viable biological material, optionally according to a predetermined pressure-time profile;
(b) keeping the said biological material at the hydrostatic pressure for a predetermined time period;
(c) releasing the hydrostatic pressure;
(d) further cryopreserving the said viable biological material using any protocol applicable thereto,
wherein the viable biological material is mammalian blastocyst or mammalian sperm.
2. The method according to claim 1 wherein the said hydrostatic pressure is applied for a time period between 1 second and 300 minutes.
3. The method according to claim 2 wherein the said hydrostatic pressure is applied for a time period between 1 second and 150 minutes.
4. The method according to claim 2 wherein the said hydrostatic pressure is applied for a time period between 1 second and 90 minutes.
5. The method according to claim 2 wherein the said hydrostatic pressure is applied for a time period between 1 second and 60 minutes.
6. The method according to claim 1 wherein the pressure is released gradually over a time period between 1 second and 4 hours.
7. The method according to claim 1 , further comprising before step (a), placing the biological material into a pressurizing device.
8. The method according to claim 7 , wherein a control system is provided for controlling the depressurization of the pressure chamber over a time period between 1 second and 4 hours.
9. The method according to claim 7 , wherein the pressurizing device includes a pressure chamber suitable for receiving the biological material, and means to provide controlled pressure in the range of 1 to 200 MPa.
10. The method according to claim 9 wherein said means provides controlled pressure in the range of 10 to 100 MPa.
11. The method according to claim 9 wherein said means provides controlled pressure in the range of 20 to 75 MPa.
12. The method according to claim 9 wherein said means provides controlled pressure in the range of 30 to 60 MPa.
13. The method according to claim 9 , wherein the pressurizing device comprises means to maintain the said pressure for a time period between 1 second and 300 minutes.
14. The method according to claim 13 , wherein said pressure is maintained for a time period between 1 second and 150 minutes.
15. The method according to claim 13 , wherein said pressure is maintained between 1 second and 60 minutes.
16. The method according to claim 1 wherein the said hydrostatic pressure is in the range of 10 to 100 MPa.
17. The method according to claim 1 , wherein the said hydrostatic pressure is in the range of 20 to 75 MPa.
18. The method according to claim 1 wherein the said hydrostatic pressure is in the range of 30 to 60 MPa.