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1. (WO2019002399) FREEZING OF BIOLOGICAL MATERIAL
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CLAIMS

1. A method of freezing human and/or animal cells in suspension, wherein one or more samples of human and/or animal cells in suspension (12) contained in container(s) (1) comprise one or more liquids, the method comprising :

determining the cooling needed such that the phase transition time of a calibration sample (11) contained in a container (1) is less than 12 minutes;

cooling said one or more samples (12) in the same manner as said calibration sample (11).

2. The method of freezing human and/or animal cells in suspension as claimed in claim 1, wherein the cooling needed is determined such that the phase transition time of a calibration sample (11) contained in a container (1) is less than 6 minutes, such as less than 3 minutes.

3. The method of freezing human and/or animal cells in suspension as claimed in claim 1 or 2, wherein the one or more samples are exposed to a magnetic field and/or a pulsing electric field during the phase transition time, for a time period of at least 1 minute, such as at least 2 minutes, such as at least 3 minutes, such as at least 4 minutes, such as the whole phase transition time.

4. The method of freezing human and/or animal cells in suspension as claimed in claim 2, wherein the strength of the magnetic field is between 0.0001 and 1.0 T.

5. The method of freezing human and/or animal cells in suspension as claimed in previous claims 2 or 3, wherein the strength of the pulsing electric field is between 0,1-lOOV/cm, and the pulsing frequency is between 1-1000 kHz.

6. The method of freezing human and/or animal cells in suspension as claimed in any previous claim, wherein said container (1) does not exceed 100 mm in any dimension, such as does not exceed 50 mm, preferably does not exceed 25 mm.

7. The method of freezing human and/or animal cells in suspension as claimed in any previous claim characterized in that a cryoprotectant is not added to said

container (1) containing said samples of human and/or animal cells in suspension (12).

8. The method of freezing human and/or animal cells in suspension as claimed in any of claims 1-6, wherein a small amount of cryoprotectant is added to the container 1 containing the biological material 12, such that less than 5% of the total sample volume is cryoprotectant, such that less than 4%, such as less than 3%, such as less than 2%, such as less than 1%.

9. The method of freezing human and/or animal cells in suspension as claimed in any previous claim, wherein the cooling of said one or more samples contained in containers (1) comprises:

placing said container (1) in a setup comprising a fan or other wind- generating means, where the wind generated cools the container (1) and the content of the container (1),

regulating the wind generating means, such as the speed of the generated wind to achieve said cooling.

10. The method of freezing human and/or animal cells in suspension as claimed in claims 1 - 4, wherein the cooling of said container (1) comprises:

placing said container (1) in a cooling bath (13),

regulating the cooling power of the cooling bath by reducing or increasing the temperature of the freezing solution to achieve said cooling .

11. The method of freezing human and/or animal cells in suspension as claimed in claims 1 - 4, wherein the cooling of said container (1) comprises:

placing said container (1) on a first plate member (2),

regulating the cooling of said first plate member (2) to achieve said cooling .

12. The method of freezing human and/or animal cells in suspension as claimed in claim 16 further comprising :

a second plate member (7) closing on said first plate member (2).

13. The method of freezing human and/or animal cells in suspension as claimed in any previous claim, wherein the samples of human and/or animal cells in suspension (12) to be frozen is human cells, animal cells, cell lines, primary cells, stem cells, tissue cells, blood products, embryos, sperm, or fish eggs.

14. An apparatus for freezing samples of human and/or animal cells in

suspension, wherein the apparatus comprises a cooling system and a control system to control the cooling power of said cooling system such that a time is set, wherein the time set is the duration of latent heat removal from a calibration sample (11).

15. An apparatus for freezing samples of human and/or animal cells in

suspension, wherein the apparatus comprises

a cooling system comprising a wind-generating member, such as a fan, configured to cool said samples over a period of time, and

a control system (23) being able to determine the cooling needed such that the phase transition time of a calibration sample (1) is less than 12 minutes; and being configured for controlling the cooling system such that said samples are cooled in the same manner as the calibration sample (11).

16. An apparatus as claimed in claim 14 or 15, wherein the control system comprise temperature sensors (24) immersed in the calibration sample (11) and wherein the control system (23) determines the cooling needed based on input data from the temperature sensors (24).

17. An apparatus as claimed in any of claims 14-16, wherein the apparatus further comprise a magnetic field generating means (25), configured to expose the one or more samples to a magnetic field strength between O.OOOIT and 1.0 T.

18. An apparatus as claimed in any of claim 14-17, wherein the magnetic field generating means (25) comprise a plurality of magnets, such as permanent block magnets, arranged in an array.

19. An apparatus as claimed in any of claims 14-18, wherein the apparatus comprise a pulsing electric field generating means, configured to expose the one or more samples to a pulsing electric field with a strength in the range of 0,1- lOOV/cm, and a pulsing frequency in the range of 1-1000 kHz.

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20. An apparatus as claimed in any of claim 14-19, wherein the pulsed electric field are made from a pulsed power supply, an electrode pair and a plate capacitor arrangement comprising two parallel spaced aluminium plates.

10 21. An apparatus as claimed in any of claims 14-20, wherein the apparatus

comprise a box (26) made from materials with low or no magnetic permeability in nature, such as aluminium, plastic or mica, the box comprising three different compartments, such as drawers, wherein a top compartment (27) and a bottom compartment (28) are configured to hold the magnetic field generating means

15 (25) and a middle compartment (29) is configured for holding said samples, as well as the calibration sample (11).

22. An apparatus as claimed in any of claims 14-21, wherein the magnetic field generating means are arranged on a magnet holder (31) which can be fitted into

20 the top compartment or bottom compartment comprising a plate with indentations for the magnets

23. An apparatus as claimed in any of claims 14-22, wherein the apparatus further comprise a sample rack (30) comprising holes adapted to hold containers

25 comprising said samples and the calibration sample, such as holes having

standard cryo-tube type diameter

24. An apparatus as claimed in any of claims 14-23, wherein the sample rack (30) is adapted to fit into the middle compartment (29).

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25. An apparatus as claimed in any of claims 14-15, further comprising a first plate member (2), which can hold a sample of biological material (12), said first plate member (2) being in thermal contact with said cooling system.

26. An apparatus as claimed in any of claims 14-15 or 25, further comprising a second plate member (7), and wherein said first plate member (2) and said second plate member (7) can move in relation to each other.

5 27. An apparatus as claimed in any of claims 14-15 or 25-26, wherein the first plate member (2) and/or the second plate member (7) has one or more indentations (5), where said indentations can hold samples of human and/or animal cells in suspension (12) or a calibration sample (11).

10 28. An apparatus as claimed in any of claims 14-15 or 25-27, wherein the cooling system comprises a wind-generating member (10).

29. An apparatus as claimed in any of claims 14-15 or 25-28, wherein the cooling system comprises a cooling bath (13).

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30. An apparatus as claimed in any of claims 14-15 or 25-29, wherein the samples of human and/or animal cells in suspension to be frozen is human cells, animal cells, cell lines, primary cells, stem cells, tissue cells, blood products, embryos, sperm, or fish eggs.