Protein A crystals and cross-linked crystals and methods of use thereof
Protein A crystals and Protein A cross-linked protein crystals (CLPCs) are described. Methods of preparing and using are also disclosed.
1. A method of purifying an immunoglobulin comprising contacting said immunoglobulin using a composition comprising cross-linked crystalline (CLCP) form of Protein A.
2. The method of claim 1 , wherein said immunoglobulin is an antibody.
3. The method of claim 2 , wherein said antibody is a therapeutic antibody.
4. The method of claim 2 , wherein said antibody is a monoclonal antibody.
5. The method of claim 1 , wherein said immunoglobulin is a Fab fragment.
6. The method of claim 1 , wherein said immunoglobulin is an Fc fragment.
7. The method of claim 1 , wherein said immunoglobulin is a single chain antibody.
8. The method of claim 1 , wherein said immunoglobulin is a chimeric antibody.
9. The method of claim 1 , wherein said immunoglobulin is a fully human antibody.
10. The method of claim 1 , wherein said immunoglobulin is a humanized antibody.
11. The method of claim 1 , wherein said immunoglobulin belongs to class IgG.
12. The method of claim 1 , wherein said composition comprises cross-linked crystalline form of Protein A, wherein the composition is cross-linked with glutaraldehyde.
13. A process for the purification of immunoglobulins which comprises:
(a) mixing a medium containing immunoglobulins with a buffer solution having a pH in the range of about pH 7.0 to pH 10 and containing a combination of cations and anions to provide a buffered immunoglobulin medium;
(b) contacting said buffered immunoglobulin medium with an immobilized cross-linked crystalline (CLCP) form of Protein A adsorbent to adsorb the immunoglobulins present in said buffered immunoglobulin medium upon said immobilized CLPC adsorbent;
(c) washing the CLPC adsorbent having immunoglobulins adsorbed thereon with said buffer solution;
(d) contacting said CLPC adsorbent having immunoglobulins adsorbed thereon with a buffer solution having a pH in the range of about pH 2 to pH 6 to remove the adsorbed immunoglobulins from the CLPC adsorbent; and
(e) recovering the removed immunoglobulins in substantially pure form.
14. The process according to claim 13 , wherein the contacting of said buffered immunoglobulin medium with said CLPC adsorbent is accomplished in a column of said CLPC adsorbent.
15. The process according to claim 13 , wherein said medium containing immunoglobulins is a normal mammalian serum.
16. The process according to claim 13 , wherein said medium containing immunoglobulins is an immune mammalian serum.
17. The process according to claim 13 , wherein said medium is a mammalian plasma.
18. The process according to claim 13 , wherein said medium is mammalian ascites fluid.
19. The process according to claim 13 , wherein said medium is obtained from a hybridoma.
20. The process according to claim 13 , wherein said medium is a tissue culture fluid.
21. The process according to claim 13 , wherein said medium is a cell culture fluid.
22. The process according to claim 13 , wherein said medium is a mammalian cell culture fluid.
23. The process according to claim 13 , wherein said medium is a bacterial cell culture fluid.
24. The process according to claim 13 , wherein said medium is a transgenic source fluid.
25. The process according to claim 13 , wherein said medium is a plant extract containing immunoglobulins.
26. The process according to claim 13 , wherein said medium is a yeast culture fluid.
27. A method of purification comprising contacting the substance to be purified with a composition comprising cross-linked crystalline form of Protein A.
28. The method of claim 1 , wherein Protein A within said CLPC is Protein A polypeptide or polypeptide fragments of Protein A.
29. The method of claim 28 , wherein Protein A and fragments thereof bind at least one immunoglobulin at the Fc-part.
30. The method of claim 1 wherein the Protein A is derived from Staphylococcus sp.
31. The method of claim 30 , wherein said Staphylococcus is Staphylococcus aureus or Staphylococcus epidermidis.
32. The method of claim 1 , wherein the cross-linking of the Protein A crystals comprises covalent links between the constituent protein molecules of the crystal.
33. The method of claim 30 , wherein the cross-linking of the Protein A crystals is through lysine amine groups, thiol groups and carbohydrate moieties.
34. The process according to claim 13 , wherein, wherein said CLPC is insoluble in said medium.