Materials and procedures for the purification of cells

• US 7,598,081 B2
• Filed: 12/20/2005
• Issued: 10/06/2009
• Est. Priority Date: 11/08/1996
• Status: Expired due to Fees

• Current Assignee: Morphogenesis, Inc. (RadioIO, Inc.)
• Original Assignee: Morphogenesis, Inc. (RadioIO, Inc.)
• Inventors: Lawman, Michael, Lawman, Patricia
• Primary Examiner(s): Saunders; David A
• Application Number:
  US11/313,058
• Publication Number:
  US 20060105399A1
• Time in Patent Office: 1,386 Days
• Field of Search: 204/403.1, 435/2, 435/7.1, 435/7.21, 435/7.24, 435/7.5, 435/30, 435/325, 435/287.2, 435/372, 436/528, 436/535, 436/806
• US Class Current: 435/325
• CPC Class Codes: C07K 16/2896 against molecules with a "CD"-designation, not provided for elsewhere
• CPC Class Codes: C07K 17/08 the carrier being a synthetic polymer
• CPC Class Codes: C12N 5/0087 Purging against subsets of blood cells, e.g. purging alloreactive T cells
• CPC Class Codes: C12N 5/0647 Haematopoietic stem cells; Uncommitted or multipotent progenitors
• CPC Class Codes: Y10S 436/806 Electrical property or magnetic property

Abstract

The subject invention provides new materials and methods for the efficient isolation and purification of stem cells. Specifically, conductive immunopolymers with stem cell specific antibodies can be used to remove stem cells from biological fluids.

Claims

1. A method for isolating target cells from a sample, said method comprising

i) contacting said sample with a conductive immunopolymer matrix, wherein said conductive immunopolymer matrix comprises:

a) a first immunopolymer layer comprising a polyheteroaromatic polymer and an Fc receptor immobilized within said polymer, wherein a first antibody is bound to said Fc receptor in said first polymer and said first antibody binds to a molecule expressed on a first target cell; and

b) a second immunopolymer layer comprising a polyheteroaromatic polymer and an Fc receptor immobilized within said polymer, wherein a second antibody is bound to said Fc receptor in said second polymer and said second antibody binds to a molecule expressed on a second target cell; and

wherein said first and second immunopolymer layers further comprise a cell monitoring system for monitoring cell binding by said first or second antibody or cell release from said first or second antibody, wherein said cell monitoring system comprises horseradish peroxidase and glucose oxidase entrapped within said first and second immunopolymers;

ii) removing said sample from contact with said conductive immunopolymer matrix, wherein said first target cell is bound to said first antibody in said first immunopolymer layer and said second target cell is bound to said second antibody in said second immunopolymer layer, thereby isolating said target cells from said sample; and

iii) washing said conductive immunopolymer matrix to remove unbound cells.

2. The method according to claim 1, wherein said antibody bound to said Fc receptor of said first immunopolymer layerbinds to a CD3, CD4, CD7, CD8, CD10, CD11b, CD14, CD19, CD20, or CD33 determinant.

3. The method according to claim 1, wherein said antibody bound to said Fc receptor of said second immunopolymer layer binds to a CD34 determinant.

4. The method according to claim 1, wherein said polyheteroaromatic polymer is an alkyl substituted polythiophene or a polypyrrole.

5. The method according to claim 1, wherein said polyheteroaromatic polymer is a naphthalene sulfonate-doped polypyrrole or a p-toluene sulfonate-doped polypyrrole.

6. The method according to claim 1, wherein said polyheteroaromatic polymer has a net negative charge at the surface of said polymer.

7. The method according to claim 1, wherein said conductive immunopolymer matrix is provided in a capillary, spiral sheet, or parallel sheet.

8. The method according to claim 1, wherein said first and second immunopolymer layers are sequentially oriented with respect to each other.

9. The method according to claim 1, wherein said target cell is a tumor cell, a stem cell, a blood cell, or a bone marrow cell.

10. The method according to claim 1, wherein said method further comprises releasing said first or second antibody from said first or second immunopolymer layer after step (iii).

11. A method for isolating target cells from a sample, said method comprising

i) contacting said sample with a conductive immunopolymer matrix, wherein said conductive immunopolymer matrix comprises:

a) a first immunopolymer layer comprising a polyphenol polymer and an Fc receptor immobilized within said polymer, wherein a first antibody is bound to said Fc receptor in said first polymer; and

said first antibody binds to a molecule expressed on a first target cell; and

b) a second immunopolymer layer comprising a polyphenol polymer and an Fc receptor immobilized within said polymer, wherein a second antibody is bound to said Fc receptor in said second polymer and said second antibody binds to a molecule expressed on a second target cell; and

wherein said first and second immunopolymer layers further comprise a cell monitoring system for monitoring cell binding by said first or second antibody or cell release from said first or second antibody, wherein said cell monitoring system comprises horseradish peroxidase and glucose oxidase entrapped within said first and second immunopolymers;

ii) removing said sample from contact with said conductive immunopolymer matrix, wherein said first target cell is bound to said first antibody in said first immunopolymer layer and said second target cell is bound to said second antibody in said second immunopolymer layer, thereby isolating said target cells from said sample; and

iii) washing said conductive immunopolymer matrix to remove unbound cells.

12. The method according to claim 11, wherein said antibody bound to said Fc receptor of said first immunopolymer layer binds to a CD3, CD4, CD7, CD8, CD10, CD11b, CD14, CD19, CD20, or CD33 determinant.

13. The method according to claim 11, wherein said antibody bound to said Fc receptor of said second immunopolymer layer binds to a CD34 determinant.

14. The method according to claim 11, wherein said polyphenol polymer has a net negative charge at the surface of said polymer.

15. The method according to claim 11, wherein said conductive immunopolymer matrix is provided in a capillary, spiral sheet, or parallel sheet.

16. The method according to claim 11, wherein said first and second immunopolymer layers are sequentially oriented with respect to each other.

17. The method according to claim 11, wherein said target cell is a tumor cell, a stem cell, a blood cell, or a bone marrow cell.

18. The method according to claim 11, wherein said method further comprises releasing said first or second antibody from said first or second immunopolymer layer after step (iii).

19. A method for isolating a target cell from a sample, said method comprising

i) contacting said sample with a conductive immunopolymer matrix, wherein said conductive immunopolymer matrix comprises:

a) an immunopolymer layer comprising a polyheteroaromatic polymer or a polyphenol polymer and an Fc receptor immobilized within said polymer, wherein an antibody is bound to said Fc receptor in said polymer and said antibody binds to a molecule expressed on said target cell; and

wherein said immunopolymer layer further comprises a cell monitoring system for monitoring cell binding by said antibody or cell release from said antibody, wherein said cell monitoring system comprises horseradish peroxidase and glucose oxidase entrapped within said immunopolymer;

ii) removing said sample from contact with said conductive immunopolymer matrix, wherein said target cell is bound to said first antibody in said first immunopolymer layer, thereby isolating said target cell from said sample; and

iii) washing said conductive immunopolymer matrix to remove unbound cells.

20. The method according to claim 19, wherein said antibody bound to said Fc receptor of said immunopolymer layerbinds to a CD3, CD4, CD7, CD8, CD10, CD11b, CD14, CD19, CD20, CD33, or CD34 determinant.

21. The method according to claim 19, wherein said antibody bound to said Fc receptor of said immunopolymer layer binds to a CD34 determinant.

22. The method according to claim 19, wherein said polyheteroaromatic polymer is an alkyl substituted polythiophene or a polypyrrole.

23. The method according to claim 19, wherein said polyheteroaromatic polymer is a naphthalene sulfonate-doped polypyrrole or a p-toluene sulfonate-doped polypyrrole.

24. The method according to claim 19, wherein said polyheteroaromatic polymer or said polyphenol polymer has a net negative charge at the surface of said polymer.

25. The method according to claim 19, wherein said conductive immunopolymer matrix is provided in a capillary, spiral sheet, or parallel sheet.

26. The method according to claim 19, wherein said target cell is a tumor cell, a stem cell, a blood cell, or a bone marrow cell.

27. The method according to claim 19, wherein said method further comprises releasing said antibody from said immunopolymer layer after step (iii).