Method for using ligands in particle separation

• US 6,153,113 A
• Filed: 02/22/1999
• Issued: 11/28/2000
• Est. Priority Date: 02/22/1999
• Status: Expired due to Fees

• Current Assignee: Caridianbct Biotechnologies LLC (Baxter International Inc.)
• Original Assignee: COBE Laboratories Incorporated (Terumo Corporation)
• Inventors: Goodrich, Raymond P., Green, Todd Curtis
• Primary Examiner(s): KIM, SUN U
• Application Number:
  US09/253,696
• Time in Patent Office: 645 Days
• Field of Search: 210/782, 210/787, 210/786, 210/789, 210/739, 435/2, 435/174, 494/37, 436/523, 436/531, 436/532
• US Class Current: 210/782
• CPC Class Codes: B01D 21/01 using flocculating agents for purifying water C02F1/52; for liquid radioactive waste G21F9/10
• CPC Class Codes: B01D 21/26 Separation of sediment aided by centrifugal force or centripetal forcecentrifuges B04B; cyclones B04C
• CPC Class Codes: B01D 21/262 by using a centrifuge
• CPC Class Codes: B01D 2221/10 Separation devices for use in medical, pharmaceutical or laboratory applications, e.g. separating amalgam from dental treatment residues
• CPC Class Codes: B03D 3/02 Coagulation
• CPC Class Codes: B04B 2005/0471 with additional elutriation separation of different particles
• CPC Class Codes: G01N 15/042 by centrifuging and investigating centrifugates centrifuges per se B04B
• CPC Class Codes: G01N 33/54313 the carrier being characterised by its particulate form

Abstract

A system and method are disclosed for separating particles having different sedimentation velocities. The system includes a container containing a binding substance including first particles and ligands attached to the first particles. When the binding substance is mixed with a liquid carrying at least second and third particles, the ligands bind the first and second particles together to form groups of bound particles. The groups of bound particles are separated from the third particles in a fluid chamber configured to be mounted on a centrifuge rotor. One of the disclosed methods includes forming a saturated fluidized bed of particles to retain the particles groups in the fluid chamber. Another of the disclosed methods includes removing at least some of the first particles after the binding substance is mixed with the liquid carrying second and third particles.

Claims

1. A method of separating particles, comprising:

mixing a binding substance including first particles and ligands attached to the first particles with a liquid carrying at least second and third particles to form a mixture, at least some of the ligands binding with the second particles to form groups of bound particles including the first and second particles;

removing a portion of the binding substance from the mixture, the portion of the binding substance removed from the mixture including first particles having attached ligands lacking binding to the second particles;

rotating a fluid chamber about an axis of rotation;

flowing at least the liquid, third particles, and groups of bound particles into the fluid chamber;

separating in the fluid chamber the third particles and the groups of bound particles according to differences in sedimentation velocity; and

allowing at least the liquid and at least a portion of the third particles to flow from the fluid chamber.

2. The method of claim 1, wherein the separating includes forming a saturated fluidized bed of particles in the fluid chamber, the saturated fluidized bed of particles retaining groups of bound particles in the fluid chamber.

3. The method of claim 2, wherein the particles forming the saturated fluidized bed include at least one of red blood cells and beads having a predetermined size.

4. The method of claim 2, wherein the particles forming the saturated fluidized bed include at least the third particles.

5. The method of claim 1, wherein the separating includes separating the third particles and the groups of bound particles by elutriation.

6. The method of claim 1, further comprising allowing incubation of the mixture to increase the binding of the ligands with the second particles.

7. The method of claim 6, further comprising agitating the mixture during the incubation to further increase the binding of the ligands with the second particles.

8. The method of claim 1, wherein the removing includes removing some of the groups of bound particles from the mixture.

9. The method of claim 1, wherein the removing includes allowing gravity sedimentation in the mixture of at least the first particles having attached ligands lacking binding to the second particles, and separating at least the sediment from the remainder of the mixture.

10. The method of claim 9, wherein the method further comprises adding a diluting medium to the mixture to facilitate the gravity sedimentation.

11. The method of claim 9, wherein the separating of the sediment from the remainder of the mixture includes decanting the remainder of the mixture from the sediment.

12. The method of claim 1, wherein the removing includes centrifugally separating at least the portion of the binding substance from the remainder of the mixture on a centrifuge.

13. The method of claim 12, wherein the method further comprises adding a diluting medium to the mixture to facilitate the centrifugal separation.

14. The method of claim 1, wherein the removing includes passing the mixture through a porous medium to filter at least the portion of the binding substance from the remainder of the mixture.

15. The method of claim 1, wherein the liquid carrying at least the second and third particles includes at least one of the group consisting of:

whole blood, blood components, a peripheral blood cell collection, a bone marrow blood cell collection, and blood components removed from an umbilical cord.

16. The method of claim 1, wherein the liquid includes plasma.

17. The method of claim 1, wherein the ligands include at least one of the group consisting of:

a protein, an antibody, a chemical composition, and mixtures thereof.

18. The method of claim 1, wherein the ligands include at least one antibody selected from the group consisting of:

anti-CD2, anti-CD3, anti-CD4, anti-CD8, anti-CD9, anti-CD10, anti-CD14, anti-CD15, anti-CD19, anti-CD20, anti-CD34, anti-CD38, anti-CMRF-44, anti-CD45, anti-CD56, anti-CD83, anti-glycophorin, anti-cytokeratin, anti-EPCAM, and combinations thereof.

19. The method of claim 1, wherein the ligands include at least one protein selected from the group consisting of:

albumin and immunoglobulin.

20. The method of claim 1, wherein the ligands include at least one of the group consisting of:

poloxamers and poly-lysine.

21. The method of claim 1, wherein an exterior surface of the first particles includes at least one of the group consisting of a surface modifying agent and poloxamer.

22. The method of claim 21, wherein the exterior surface of the first particles includes a surface modifying agent coomprising a polylactone-polysiloxane-polylactone triblock copolymer.

23. The method of claim 1, wherein the first particles are rigid, generally spherical beads.

24. The method of claim 1, wherein the second particles include at least one of the group consisting of:

antigen-specific white blood cells, CD2+ cells, CD3+ cells, CD4+ cells, CD8+ cells, CD9+ cells, CD10+ cells, CD14+ cells, CD15+ cells, CD19+ cells, CD20+ cells, CD34+ cells, CD38+ cells, CMRF-44+ cells, CD45+ cells, CD56+ cells, CD83+ cells, glycophorin+ cells, cytokeratin+ cells, EPCAM+ cells, and combinations thereof.

25. The method of claim 1, wherein the third particles include at least one of the group consisting of:

red blood cells, antigen-specific white blood cells, platelets, proteins, drugs, cytokines, and combinations thereof.

26. The method of claim 1, wherein the second and third particles include different types of antigen-specific white blood cells.

27. The method of claim 1, further comprising

passing into the fluid chamber a diluting liquid, the diluting liquid having a density less than that of the first, second, and third particles so that the diluting liquid reduces an overall density of substances in the fluid chamber and thereby limits Coriolis jetting of at least one of the liquid carrying the particles and the diluting liquid in the fluid chamber;

and

allowing the diluting liquid to flow from the fluid chamber.

28. The method of claim 27, further comprising mixing the liquid carrying particles and the diluting liquid so that at least portions of the passing and flowing are performed simultaneously.

29. The method of claim 1, further comprising flowing the liquid and third particles from the fluid chamber into a separation vessel, separating the third particles from the liquid in the separation vessel, and collecting at least the third particles in a container.

30. The method of claim 1, further comprising adding an adhesion-reducing agent to the liquid carrying particles.

31. The method of claim 1 wherein the first particles include a dye, and wherein the method further comprises monitoring substances flowing from an outlet of the fluid chamber to detect the dye.

32. The method of claim 1, further comprising monitoring substances flowing from an outlet of the fluid chamber to detect first particles flowing from the outlet, and removing the first particles from the substances.

33. A method of separating particles, comprising:

mixing a binding substance including first particles and ligands attached to the first particles with a liquid carrying at least second and third particles to form a mixture, at least some of the ligands binding with the second particles to form groups of bound particles including the first and second particles;

rotating a fluid chamber about an axis of rotation;

flowing at least the liquid, third particles, and groups of bound particles into the fluid chamber;

forming, in the fluid chamber, a saturated fluidized bed of particles, the saturated fluidized bed of particles retaining the groups of bound particles in the fluid chamber; and

allowing at least the liquid and at least a portion of the third particles to flow from the fluid chamber.

34. The method of claim 33, wherein the particles forming the saturated fluidized bed include at least one of red blood cells and beads having a predetermined size.

35. The method of claim 33, wherein the particles forming the saturated fluidized bed include at least the third particles.

36. The method of claim 33, further comprising allowing incubation of the mixture to increase the binding of the ligands with the second particles.

37. The method of claim 33, further comprising agitating the mixture during the incubation to further increase the binding of the ligands with the second particles.

38. The method of claim 33, wherein the liquid carrying at least the second and third particles includes at least one of the group consisting of:

whole blood, blood components, a peripheral blood cell collection, a bone marrow blood cell collection, and blood components removed from an umbilical cord.

39. The method of claim 33 wherein the liquid includes plasma.

40. The method of claim 33, wherein the ligands include at least one of the group consisting of:

a protein, an antibody, a chemical composition, and mixtures thereof.

41. The method of claim 33, wherein the ligands include at least one antibody selected from the group consisting of:

anti-CD2, anti-CD3, anti-CD4, anti-CD8, anti-CD9, anti-CD10, anti-CD14, anti-CD15, anti-CD19, anti-CD20, anti-CD34, anti-CD38, anti-CMRF-44, anti-CD45, anti-CD56, anti-CD93, anti-glycophorin, anti-cytokertin, anti-EPCAM, and combinations thereof.

42. The method of claim 33, wherein the ligands include at least one protein selected from the group consisting of:

albumin and immunoglobulin.

43. The method of claim 33, wherein the ligands include at least one of the group consisting of:

poloxamers and poly-lysine.

44. The method of claim 33, wherein an exterior surface of the first particles includes at least one of the group consisting of a surface modifying agent and poloxamer.

45. The method of claim 44, wherein the exterior surface of the first particles includes a surface modifying agent coomprising a polylactone-polysiloxane-polylactone triblock copolymer.

46. The method of claim 33, wherein the first particles are rigid, generally spherical beads.

47. The method of claim 33, wherein the second particles include at least one of the group consisting of:

antigen-specific white blood cells, CD2+ cells, CD3+ cells, CD4+ cells, CD8+ cells, CD9+ cells, CD10+ cells, CD14+ cells, CD15+ cells, CD19+ cells, CD20+ cells, CD34+ cells, CD38+ cells, CMRF-44+ cells, CD45+ cells, CD56+ cells, CD83+ cells, glycophorin+ cells, cytokeratin+ cells, EPCAM+ cells, and combinations thereof.

48. The method of claim 33, wherein the third particles include at least one of the group consisting of:

red blood cells, antigen-specific white blood cells, platelets, proteins, drugs, cytokines, and combinations thereof.

49. The method of claim 33, wherein the second and third particles include different types of antigen-specific white blood cells.

50. The method of claim 33, wherein the liquid carries antigen-specific white blood cells, the saturated fluidized bed of particles permitting at least some of the antigen-specific white blood cells to flow from the fluid chamber.

51. The method of claim 33, further comprising

passing into the fluid chamber a diluting liquid, the diluting liquid having a density less than that of the first, second, and third particles so that the diluting liquid reduces an overall density of substances in the fluid chamber and thereby limits Coriolis jetting of at least one of the liquid carrying the particles and the diluting liquid in the fluid chamber;

and

allowing the diluting liquid to flow from the fluid chamber.

52. The method of claim 51, further comprising mixing the liquid carrying particles and the diluting liquid so that at least portions of the passing and flowing are performed simultaneously.

53. The method of claim 33, further comprising flowing the liquid and third particles from the fluid chamber into a separation vessel, separating the third particles from the liquid in the separation vessel, and collecting at least the third particles in a container.

54. The method of claim 33, further comprising adding an adhesion-reducing agent to the liquid carrying particles.

55. The method of claim 33, wherein the first particles include a dye, and wherein the method further comprises monitoring substances flowing from an outlet of the fluid chamber to detect the dye.

56. The method of claim 33, further comprising monitoring substances flowing from an outlet of the fluid chamber to detect first particles flowing from the outlet, and removing the first particles from the substances.