Use of a filter for defrosting cells
The invention relates to a method for cell defrosting, comprising a step of defrosting a cell suspension consisting of a freezing medium and of cells, followed by a step of removing said freezing medium by filtration, said step of removing the freezing medium being carried out without the previous or simultaneous supply of medium. In particular, the filter is hydrophobic or, before addition of the cell suspension to be frozen, has been covered with a hydrophobic liquid forming a temporary barrier between the filter and the cell suspension. During defrosting, said hydrophobic liquid is removed by the application of a driving force which makes the liquid pass through the filter.
1. A cell thawing kit comprising at least one tube of frozen cells and one device equipped with a filter, the pores of which have an average diameter of between 1 and 15 microns, said device being sterile.
2. The kit as claimed in claim 1 , wherein the cells have been frozen in the device equipped with a filter.
3. The kit as claimed in claim 1 , wherein the cells are VERO cells.
4. The kit as claimed in claim 1 , wherein the device equipped with a filter is a centrifugal filter.
5. The kit as claimed in claim 1 , wherein the kit contains at least two sterile centrifugal filters.
6. The kit as claimed in claim 1 , wherein the average diameter of the pores of the filter is between 2 and 10 microns.
7. The kit as claimed in claim 1 , for promoting the embryo culture or oocyte in vitro maturation, wherein the kit also comprises one or more of the following elements: 200 μl and/or 1000 μl sterile tips, a culture support, sterile tubes, and sterile pipettes.
8. The kit as claimed in claim 1 , for promoting the embryo culture or oocyte in vitro maturation, wherein the kit also comprises a rinsing medium and/or an appropriate culture medium.
9. The kit as claimed in claim 1 , further comprising
a container having hermetic closing means capable of containing the device equipped with a filter,
wherein the filter has hydrophobic properties.
10. The kit as claimed in claim 9 , wherein the hermetic closing means comprise a screw cap.
11. A cell thawing method using the cell thawing kit of claim 1 , comprising
a step of thawing a cell suspension consisting of a freezing medium and said frozen cells, followed by a step of removing said freezing medium by filtration using said device equipped with a filter, said step of removing the freezing medium being carried out without prior or simultaneous provision of medium.
12. The method as claimed in claim 11 , comprising the following steps:
(i) placing a first container, containing a frozen cell suspension, at a temperature which allows the cell suspension to thaw;
(ii) transferring the cell suspension from the first container into the device equipped with a filter;
(iii) applying to the cell suspension a force which drives the passing of the freezing medium through the filter; and
(iv) resuspending the cells in a solution.
13. The method as claimed in claim 11 , comprising the following steps:
(i) placing a first container, containing a frozen cell suspension in the device equipped with a filter, at a temperature which allows the cell suspension to thaw;
(ii) applying to the cell suspension a force which drives the passing of the freezing medium through the filter; and
(iii) resuspending the cells in a solution.
14. The method as claimed in claim 12 , wherein the device equipped with a filter is a centrifugal filter.
15. The method as claimed in claim 12 , wherein the average diameter of the pores of the filter is between 2 and 10 microns.
16. The method as claimed in claim 11 , wherein the passing of the freezing medium through the filter is obtained by centrifuging the device equipped with a filter, placed in a second container.
17. The method as claimed in claim 16 , wherein the device is centrifuged for a period of less than or equal to 10 minutes at a speed which makes it possible to apply to the cell suspension an acceleration of between 50 and 1500 g, preferably between 100 and 200 g.
18. The method as claimed in claim 12 , wherein the passing of the freezing medium through the filter is obtained by applying an increased pressure on the side of the filter containing the cell suspension, and/or a reduced pressure on the other side of the filter.
19. The method as claimed in claim 12 , wherein the amount of cells placed in each device equipped with a filter is between 10 4 and 10 7 .
20. The method as claimed in claim 11 , wherein the step of resuspending the cells is carried out by performing pipetting operations using a P200 or P 1000 pipette.
21. The method as claimed in claim 13 , wherein the filter is hydrophobic, or, before the addition of the cell suspension to be frozen, has been covered with a hydrophobic liquid forming a temporary barrier between the filter and the cell suspension, said liquid being removed in step (ii) during the application of the driving force.