Microfluidic Detection of Analytes

• US 20080202933A1
• Filed: 10/04/2006
• Published: 08/28/2008
• Est. Priority Date: 10/04/2005
• Status: Active Grant

• Current Assignee: Headway Technologies Incorporated (TDK Corporation)
• Original Assignee: Headway Technologies Incorporated (TDK Corporation)
• Inventors: Hu, Celine
• Granted Patent: US 8,263,022 B2
• Time in Patent Office: Days
• US Class Current: 204/451
• CPC Class Codes: B01L 2200/027 for microfluidic devices
• CPC Class Codes: B01L 2300/0636 Integrated biosensor, microarrays
• CPC Class Codes: B01L 2400/0421 electrophoretic flow
• CPC Class Codes: B01L 3/502715 characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
• CPC Class Codes: B33Y 80/00 Products made by additive manufacturing
• CPC Class Codes: G01N 1/40 Concentrating samples
• CPC Class Codes: G01N 2001/4038 electric methods, e.g. electromigration, electrophoresis, ionisation
• CPC Class Codes: G01N 27/447 using electrophoresis
• CPC Class Codes: G01N 33/536 with immune complex formed in liquid phase

Abstract

An apparatus and methods for concentrating samples for application to microfluidic devices are disclosed. The methods involve electrophoresing charged molecules from a high volume sample into a smaller volume. The analyte of interest can be a charged molecule or can be modified to be charged using, for example, one or more ionic moieties.

Claims

1. A method for introducing an analyte of interest in a sample to a microfluidic device, comprising:

providing an aqueous sample in a large volume reservoir, said sample containing the analyte and having a volume of greater than 10 microliters, wherein the analyte of interest is charged or has been associated with a charged molecule;

providing a microfluidic device comprising an analysis area;

providing a connector, wherein the large volume reservoir and the microfluidic device are fluidically connected via the connector;

electrophoresing the analyte from the large volume reservoir to the microfluidic device via the connector; and

further transporting the analyte to the analysis area for a time sufficient to result in a higher concentration of analyte in the analysis area than the concentration in the sample.

2. The method of claim 1, further comprising providing at least one other aqueous sample in a large volume reservoir fluidically connected to the microfluidic device and further comprising at least one other analysis area for said one other sample.

3. The method of claim 1, wherein the microfluidic device comprises a staging reservoir, and wherein the large volume reservoir and the staging reservoir are fluidically connected via the connector.

4. The method of claim 1, wherein the large volume reservoir is a chamber that is not integral with said microfluidic device.

5. The method of claim 4, wherein said large volume reservoir is a well of a microwell plate, an eppendorf tube, or a test tube.

6. The method of claim 1, wherein said connector is a capillary tubing.

7. The method of claim 1, further comprising admixing an antibody into said sample before electrophoresis, wherein said antibody specifically binds to said analyte of interest.

8. The method of claim 7, wherein said antibody is modified with at least one ionic moiety or is modified to be associated with at least one ionic moiety.

9. The method of claim 8 wherein the ionic moiety is selected from the group consisting of:

nucleic acids, poly-amines, sulfated glycans and succinylated proteins, polyacrylic acid, polymethacrylic acid, polyethylacrylic acid, polypropylacrylic acid, polybutylacrylic acid, polymaleic acid, dextran sulfate, heparin, hyaluronic acid, polysulfates, polysulfonates, polyvinyl phosphoric acid, polyvinyl phosphonic acid, copolymers of polymaleic acid, polyhydroxybutyric acid and mixed polymers.

11. The method of claim 7, wherein the microfluidic device comprises a staging reservoir fluidically connected to the large volume reservoir via the connector, and wherein the method further comprising comprises removing said antibody before the analyte of interest has been transferred to said staging reservoir.

12. The method of claim 1, wherein said analyte is covalently bound to an ionic moiety or is associated with a charged molecule by being bound to a carrier molecule that is ionically charged or is modified to be ionically charged.

14. The method of claim 12, wherein the carrier molecule is selected from the group consisting of an antibody, a receptor, a ligand, and an antigen.

15. The method of claim 1, wherein said analyte of interest is a biomolecule.

16. The method of claim 15, wherein said biomolecule is selected from the group consisting of a peptide, a protein, a nucleic acid, a lipid and a sugar.

17. The method of claim 16, wherein said analyte of interest is a nucleic acid.

18. The method of claim 1, wherein the analysis area comprises a capture site comprising a capture agent, said method further comprising transporting the analyte across the capture site under conditions where the analyte of interest is captured by the capture agent.

19. The method of claim 1, wherein the analyte, when transported to the analysis area, is bound to an antibody or an ionic moiety.

20. The method of claim 19, wherein the analyte is bound to an antibody and the antibody is covalently or noncovalently bound to a charged molecule.

21. The method of claim 1, wherein the analyte, when electrophoresed from the large volume reservoir to the microfluidic device, is bound to (i)an antibody that is covalently or noncovalently bound to a charged molecule or (ii) an ionic moiety.

30. The method of claim 18, wherein said transport is by electrophoresis.

31. The method of claim 18, further comprising detecting the analyte bound by the capture agent.

36. The method of claim 1, wherein said sample has a volume of greater than about 20 μ

l.

37. The method of claim 36, wherein said sample has a volume of greater than about 50 μ

l.

38. The method of claim 31, wherein said detecting comprises detecting a label on the antibody, the analyte, or the ionic moiety.

39. The method of claim 1, further comprising immobilizing said analyte of interest on a microparticle before electrophoresis.

40. The method of claim 39, wherein said microparticle is a magnetic microparticle.

41. The method of claim 39, wherein said microparticle is coated with at least one receptor, antibody, or anti-ligand that specifically binds said analyte of interest or a group of molecules including said analyte of interest.

42. The method of claim 39, further comprising the step of removing the analyte from the microparticle prior to electrophoresis.

45. The method of claim 41, wherein the method further comprises detecting a label on a second antibody bound to said analyte of interest or said group of molecules including said analyte of interest.

56. The method of claim 19, wherein the analyte is bound to an ionic moiety selected from the group consisting of:

nucleic acids, poly-amines, sulfated glycans and succinylated proteins, polyacrylic acid, polymethacrylic acid, polyethylacrylic acid, polypropylacrylic acid, polybutylacrylic acid, polymaleic acid, dextran sulfate, heparin, hyaluronic acid, polysulfates, polysulfonates, polyvinyl phosphoric acid, polyvinyl phosphonic acid, copolymers of polymaleic acid, polyhydroxybutyric acid and mixed polymers.

57. The method of claim 56, wherein said ionic moiety is a nucleic acid.

58. The method of claim 21, wherein the analyte is bound to an ionic moiety selected from the group consisting of:

nucleic acids, poly-amines, sulfated glycans and succinylated proteins, polyacrylic acid, polymethacrylic acid, polyethylacrylic acid, polypropylacrylic acid, polybutylacrylic acid, polymaleic acid, dextran sulfate, heparin, hyaluronic acid, polysulfates, polysulfonates, polyvinyl phosphoric acid, polyvinyl phosphonic acid, copolymers of polymaleic acid, polyhydroxybutyric acid and mixed polymers.

59. The method of claim 58, wherein said ionic moiety is a nucleic acid.

60. The method of claim 19, wherein the microfluidic device comprises a staging reservoir, and wherein the large volume reservoir and the staging reservoir are fluidically connected via the connector, and further wherein the analyte is electrophoresed from the large volume reservoir to the staging reservoir.

61. The method of claim 21, wherein the microfluidic device comprises a staging reservoir, and wherein the large volume reservoir and the staging reservoir are fluidically connected via the connector, and further wherein the analyte is electrophoresed from the large volume reservoir to the staging reservoir.

10. (canceled)

13. (canceled)

22-29. -29. (canceled)

32-35. -35. (canceled)

43-44. -44. (canceled)

46. (Canceled)

47. A microfluidic device for detection of an analyte comprising:

a) a staging reservoir in fluidic communication with an analysis area that comprises capture agents for the (i) the analyte, (ii) an antibody or (iii) a nucleic acid; and

b) a first microelectrode located in the staging reservoir and a second microelectrode distal to at least a portion of the analysis area, said microelectrodes being positioned such that an electric field may be generated between the electrodes and a charged analyte electrophoresed between the electrodes.

48. A system for introducing a charged analyte in a sample to a microfluidic device, comprising:

at least one large volume reservoir, operably attached to a first electrode;

a microfluidic device having at least one analysis area, wherein said microfluidic device is operably attached to a second electrode; and

at least one connector for moving charged analytes from said large volume reservoir to said microfluidic device, wherein said microfluidic device comprises a staging reservoir and wherein said large volume reservoir is fluidically connected to said staging reservoir via the connector.

49. The system of claim 48, wherein said at least one connector is capillary tubing.

51. The system of claim 48, wherein the analysis area comprises a capture site.

54. The system of claim 48, wherein said second electrode is operably attached at the analysis area or to said staging reservoir.

50. (canceled)

52-53. -53. (canceled)

55. (canceled)