Methods for freezing and thawing proteins

Abstract

A method for freezing and thawing proteins is disclosed. The proteins are rapidly frozen by introducing droplets of a protein solution into a cryogenic freezing medium. The frozen pellets thus formed are rapidly thawed by introducing them into a heat transfer fluid to form a reconstitute protein solution.

1 . A method comprising feeding droplets to a freezing medium thereby freezing said droplets and forming pellets. 
     
     
         2 . The method as claimed in  claim 1  wherein said droplets comprise a protein. 
     
     
         3 . The method as claimed in  claim 2  wherein said protein is in solution. 
     
     
         4 . The method as claimed in  claim 1  wherein said freezing medium is a cryogen. 
     
     
         5 . The method as claimed in  claim 1  wherein said cryogen is selected from the group consisting of liquid nitrogen, oxygen, air, and argon. 
     
     
         6 . The method as claimed in  claim 1  further comprising adding stabilizers to said freezing medium. 
     
     
         7 . The method as claimed in  claim 6  wherein said stabilizers are selected from the group consisting of sorbitol, sucrose, trelose, and alenine. 
     
     
         8 . The method as claimed in  claim 1  further comprising adding bulking agents and buffers to said freezing medium. 
     
     
         9 . The method as claimed in  8  wherein said bulking agents are selected from the group consisting of glycine and mannitol and said buffers are selected from the group consisting of sodium citrate and sodium phosphate. 
     
     
         10 . The method as claimed in  claim 2  wherein said protein is pre-cooled to a temperature range of −20° C. to −45° C. 
     
     
         11 . The method as claimed in  claim 1  wherein the temperature of said freezing medium is −80° C. 
     
     
         12 . The method as claimed in  claim 1  wherein said droplets are added to said freezing medium from 0.5 to 15 seconds. 
     
     
         13 . The method as claimed in  claim 1  wherein said pellets are 0.5 to 15 mm in diameter. 
     
     
         14 . The method as claimed in  claim 1  wherein said pellets are separated from said freezing medium. 
     
     
         15 . The method as claimed in  claim 1  wherein said pellets are stored at temperatures of −80° C. or below. 
     
     
         16 . A method comprising reconstituting vitrified pellets by adding said vitrified pellets to a heat transfer fluid. 
     
     
         17 . The method as claimed in  claim 16  wherein said vitrified pellets comprise a protein solution. 
     
     
         18 . The method as claimed in  claim 16  wherein heat transfer fluid is water. 
     
     
         19 . The method as claimed in  claim 16  wherein said heat transfer fluid is in a stirred vessel. 
     
     
         20 . The method as claimed in  claim 16  wherein said vitrified pellets are added to said heat transfer fluid in an amount necessary to form a reconstituted protein solution. 
     
     
         21 . A method comprising feeding droplets to a freezing medium thereby freezing said droplets and forming pellets and reconstituting said pellets by adding said pellets to a heat transfer solution. 
     
     
         22 . The method as claimed in  claim 21  wherein said droplets comprise a protein. 
     
     
         23 . The method as claimed in  claim 22  wherein said protein is in solution. 
     
     
         24 . The method as claimed in  claim 21  wherein said freezing medium is a cryogen. 
     
     
         25 . The method as claimed in  claim 21  wherein said cryogen is selected from the group consisting of liquid nitrogen, oxygen, air, and argon. 
     
     
         26 . The method as claimed in  claim 21  further comprising adding stabilizers to said freezing medium. 
     
     
         27 . The method as claimed in  claim 26  wherein said stabilizers are selected from the group consisting of sorbitol, sucrose, trelose, and alenine. 
     
     
         28 . The method as claimed in  claim 21  further comprising adding bulking agents and buffers to said freezing medium. 
     
     
         29 . The method as claimed in  28  wherein said bulking agents are selected from the group consisting of glycine and mannitol and said buffers are selected from the group consisting of sodium citrate and sodium phosphate. 
     
     
         30 . The method as claimed in  claim 22  wherein said protein is pre-cooled to a temperature range of −20° C. to −45° C. 
     
     
         31 . The method as claimed in  claim 21  wherein the temperature of said freezing medium is −80° C. 
     
     
         32 . The method as claimed in  claim 21  wherein said droplets are added to said freezing medium from 0.5 to 15 seconds. 
     
     
         33 . The method as claimed in  claim 21  wherein said pellets are 0.5 to 15 mm in diameter. 
     
     
         34 . The method as claimed in  claim 21  wherein said pellets are separated from said freezing medium. 
     
     
         35 . The method as claimed in  claim 21  wherein said pellets are stored at temperatures of −80° C. or below. 
     
     
         36 . The method as claimed in  claim 21  wherein said vitrified pellets comprise a protein solution. 
     
     
         37 . The method as claimed in  claim 21  wherein heat transfer fluid is water. 
     
     
         38 . The method as claimed in  claim 21  wherein said heat transfer fluid is in a stirred vessel. 
     
     
         39 . The method as claimed in  claim 21  wherein said vitrified pellets are added to said heat transfer fluid in an amount necessary to form a reconstituted protein solution.