Method for freeze-drying cells in a hydrogel comprising nanofibrillar cellulose and freeze-dried cells in an aerogel comprising nanofibrillar cellulose

Patent metadata

Field	Value
Publication number	US-11324701-B2
Application number	US-201716468390-A
Country	US
Kind code	B2
Filing date	Aug 25, 2017
Priority date	Dec 15, 2016
Publication date	May 10, 2022
Grant date	May 10, 2022

Abstract

The present disclosure provides a method for freeze-drying cells in a hydrogel comprising nanofibrillar cellulose, the method comprising providing a hydrogel comprising nanofibrillar cellulose, providing cells, combining the cells and the hydrogel comprising nanofibrillar cellulose to form a cell system, and freeze drying the cell system to obtain dried cells in a hydrogel comprising nanofibrillar cellulose. The present disclosure also provides a freeze-dried hydrogel comprising nanofibrillar cellulose and cells.

First claim

The invention claimed is:

1. A method for freeze-drying cells in a hydrogel comprising nanofibrillar cellulose, the method comprising:
providing a hydrogel comprising nanofibrillar cellulose,
providing cells,
combining the cells and the hydrogel comprising nanofibrillar cellulose to form a cell system, and
freeze drying the cell system to obtain dried cells in an aerogel comprising nanofibrillar cellulose;
wherein the cells are eukaryotic cells.

2. The method of claim 1 , further comprising providing one or more cryoprotective agent(s) and/or one or more lyoprotective agent(s), and adding the one or more cryoprotective agent(s) and/or one or more lyoprotective agent(s) to the cell system.

3. The method of claim 1 , further comprising culturing the cells in the cell system.

4. The method of claim 1 , wherein the cells are mammalian cells.

5. The method of claim 4 , wherein the cells are cancer cells.

6. The method of claim 2 , wherein the one or more cryoprotective and/or lyoprotective agent(s) is selected from trehalose, glycerol, and polyethylene glycol.

7. The method of claim 2 , wherein the one or more cryoprotective and/or lyoprotective agent(s) comprise trehalose and glycerol.

8. The method of claim 1 , wherein the nanofibrillar cellulose, when dispersed in water, provides a Brookfield viscosity of at least 2000 mPa·s, measured at 20° C.±1° C., at a consistency of 0.8% (w/w) and at 10 rpm.

9. The method of claim 1 , wherein the concentration of the nanofibrillar cellulose in the hydrogel before the freeze-drying is in the range of 0.1-10% (w/w).

10. The method of claim 1 , wherein the nanofibrillar cellulose is selected from anionically modified nanofibrillar cellulose, cationically modified nanofibrillar cellulose and unmodified nanofibrillar cellulose, and TEMPO oxidized nanofibrillar cellulose.

11. The method of claim 1 , wherein the freeze drying is continued until the dried hydrogel has a moisture content 10% (w/w) or less.

12. The method of claim 1 , wherein the freeze drying comprises first lowering the temperature of the cell system to at least to −20° C., and after that applying lowered pressure to remove water from the cell system.

13. The method of claim 1 , wherein the nanofibrillar cellulose has a number average fibril diameter of 1 to 100 nanometers.

14. The method of claim 1 , wherein the nanofibrillar cellulose, when dispersed in water, provides a zero-shear viscosity of 5000-50000 Pa·s, and a yield stress of 3-15 Pa, determined by rotational rheometer at a consistency of 0.5% (w/w) in aqueous medium at 22° C.

15. A freeze-dried aerogel comprising freeze-dried nanofibrillar cellulose and freeze-dried cells, wherein the moisture content of the freeze-dried aerogel is 10% or less, wherein the freeze-dried cells are eukaryotic cells.

16. The freeze-dried aerogel of claim 15 , further comprising one or more cryoprotective agent(s) and/or one or more lyoprotective agent(s).

17. The freeze-dried aerogel of claim 15 , wherein the freeze-dried cells are mammalian cells.

18. The freeze-dried aerogel of claim 17 , wherein the cells are cancer cells.

19. The freeze-dried aerogel of claim 16 , wherein the one or more cryoprotective and/or lyoprotective agent(s) is selected from trehalose, glycerol, and polyethylene glycol.

20. The freeze-dried aerogel of claim 16 , wherein the one or more cryoprotective and/or lyoprotective agent(s) agents comprise trehalose and glycerol.

21. The freeze-dried aerogel of claim 15 , wherein the content of the cells in the freeze-dried aerogel is in the range of 0.1-65% (w/w).

22. The freeze-dried aerogel of claim 16 , wherein the content of the one or more cryoprotective and/or lyoprotective agent(s) is in the range of 1-10% (w/w) and/or the content of the trehalose is in the range of 0.5-50% (w/w) in the freeze-dried aerogel.

23. The freeze-dried aerogel of claim 15 , wherein the nanofibrillar cellulose, when dispersed in water, provides a Brookfield viscosity of at least 2000 mPa·s, measured at 20° C.±1° C., at a consistency of 0.8% (w/w) and at 10 rpm.

24. The freeze-dried aerogel of claim 15 , wherein the nanofibrillar cellulose is selected from anionically modified nanofibrillar cellulose, cationically modified nanofibrillar cellulose and unmodified nanofibrillar cellulose, and TEMPO oxidized nanofibrillar cellulose.

25. The freeze-dried aerogel of claim 15 , obtained with the method of claim 1 .

26. The freeze-dried aerogel of claim 15 , wherein the freeze-dried nanofibrillar cellulose has a number average fibril diameter of 1 to 100 nanometers.

27. The freeze-dried aerogel of claim 15 , wherein the nanofibrillar cellulose, when dispersed in water, provides a zero-shear viscosity of 5000-50000 Pa·s, and a yield stress of 3-15 Pa, determined by rotational rheometer at a consistency of 0.5% (w/w) in aqueous medium at 22° C.

Assignees

• Upm Kymmene Corp

Inventors

• Paukkonen Heli
• Auvinen Vili-Veli
• Yliperttula Marjo
• Laurén Patrick
• Urtti Arto
• Laaksonen Timo
• Merivaara Arto
• Hakkarainen Tiina
• Monni Outi
• Mäkelä Anne
• Somersalo Petter
• Karhemo Piia-Riitta
• Joensuu Heikki
• Koivuniemi Raili
• Luukko Kari
• Nuopponen Markus

Classifications

• A01N1/128

  Chemically defined matrices for immobilising, holding or storing living parts, e.g. alginate gels; Chemically altering living parts, e.g. by cross-linking · CPC title

• A01N1/125

  Freeze protecting agents, e.g. cryoprotectants or osmolarity regulators · CPC title

• A61L26/008

  Hydrogels or hydrocolloids · CPC title

• F26B5/06

  the process involving freezing · CPC title

• A61L2400/12

  Nanosized materials, e.g. nanofibres, nanoparticles, nanowires, nanotubes; Nanostructured surfaces · CPC title