METHODS AND APPARATUS FOR MAGNETIC MULTI-BEAD ASSAYS

• US 20190331674A1
• Filed: 12/22/2017
• Published: 10/31/2019
• Est. Priority Date: 12/23/2016
• Status: Active Grant

• Current Assignee: Quantum Diamond Technologies Inc.
• Original Assignee: Quantum Diamond Technologies Inc.
• Inventors: Connolly, Colin B., Randall, Jeffrey D., Pena, John C.
• Granted Patent: US 11,513,115 B2
• Time in Patent Office: Days
• CPC Class Codes: G01N 24/10 by using electron paramagnetic resonance G01N24/12 takes precedence
• CPC Class Codes: G01N 27/745 for detecting magnetic beads used in biochemical assays concerning the assays G01N33/54326; sensors therefor G01R33/1269; automatic analysers therefor G01N35/0098
• CPC Class Codes: G01N 33/536 with immune complex formed in liquid phase
• CPC Class Codes: G01N 33/54326 Magnetic particles
• CPC Class Codes: G01N 33/553 Metal or metal coated
• CPC Class Codes: G01R 33/26 using optical pumping
• CPC Class Codes: G01R 33/323 Detection of MR without the use of RF or microwaves, e.g. force-detected MR, thermally detected MR, MR detection via electrical conductivity, optically detected MR

Abstract

The present application discloses methods and apparatus for detecting a complex including an analyte that include contacting a sample in a solution with a population of functionalized beads of a first type, which are magnetic functionalized beads and are functionalized to include a first moiety that associates with an analyte under suitable conditions, contacting the sample solution with a population of functionalized beads of a second type, which are functionalized to include a second moiety that associates with the analyte under suitable conditions, contact resulting in formation of a complex including one of the first type of functionalized bead, the analyte, and one of the second type of functionalized bead, and detecting the complex including the analyte by detecting magnetic fields produced by the magnetic functionalized bead and by detecting the functionalized bead of the second type associated with the analyte in the complex.

Claims

1. A bead-based magnetic assay system for detecting a complex including an analyte based on optically detected magnetic resonance (ODMR), the system comprising:

(a) a plurality of functionalized beads of a first type, which are magnetic functionalized beads and are functionalized to include a first moiety that associates with an analyte under suitable conditions;

(b) a plurality of functionalized beads of a second type, which are functionalized to include a second moiety that associates with the analyte under suitable conditions;

(c) a substrate including at least one ODMR center;

(d) a light source configured to generate incident light that excites electrons within the at least one ODMR center from a ground state to an excited state;

(e) a magnet for applying a bias magnetic field on a complex disposed over the at least one ODMR center, the complex including one of the first type of functionalized bead, the analyte, and one of the second type of functionalized bead;

(f) a microwave source configured to generate a microwave field incident on the at least one ODMR center, the microwave source being further configured to generate the microwave field with frequencies which correspond to ground state transitions in the at least one ODMR center, in which the at least one ODMR center produces emitted light when illuminated by the incident light, characteristics of the emitted light being influenced by the microwave field and by the magnetic functionalized bead associated with the analyte in the complex; and

(g) an optical photodetector that detects light emitted by the at least one ODMR center.

2. The system of claim 1, wherein the at least one ODMR center is a silicon vacancy center in a silicon carbide lattice, or in a diamond lattice.

4. The system of claim 1, wherein the at least one ODMR center is a nitrogen-vacancy center in a diamond lattice.

5. The system of claim 4, wherein the at least one ODMR center is formed in an upper surface of the substrate, or the at least one ODMR center is a plurality of ODMR centers formed in the upper surface of the diamond substrate.

7. The system of claim 5, wherein the optical photodetector is an optical imaging system having an imaging sensor that images the emitted light from the plurality of ODMR centers.

8. The system of claim 1, wherein each of the first and the second moiety is a receptor, protein, antibody, cell, virus, or nucleic acid sequence.

9. The system of claim 1, wherein the functionalized beads of the first type are superparamagnetic functionalized beads including a superparamagnetic material.

10. The system of claim 9, wherein the functionalized beads of the first type include a nonmagnetic layer encapsulating the superparamagnetic material.

11. The system of claim 9, wherein the superparamagnetic functionalized beads include iron oxide particles.

12. The system of claim 1, wherein the functionalized beads of the first type comprise magnetic nanoparticles disposed within a polymer substrate, or on a surface of the polymer substrate.

14. The system of claim 1, wherein the functionalized beads of the second type are fluorescent functionalized beads.

15. The system of claim 1, wherein the functionalized beads of the second type are magnetic functionalized beads including a quantity of magnetic material distinguishable from the functionalized beads of the first type.

16. The system of claim 1, wherein the functionalized beads of the second type are magnetic functionalized beads, the second type of functionalized beads including a magnetic property distinguishable from the functionalized beads of the first type.

17. The system of claim 16, wherein the functionalized beads of the first type are superparamagnetic functionalized beads including a superparamagnetic material.

18. The system of claim 17, wherein the functionalized beads of the first type include a nonmagnetic layer encapsulating the superparamagnetic material.

19. The system of claim 16, wherein the functionalized beads of the second type are ferromagnetic functionalized beads including a ferromagnetic material.

20. The system of claim 19, wherein the functionalized beads of the second type include a nonmagnetic layer encapsulating the ferromagnetic material.

21. The system of claim 1, wherein each of the first type of functionalized beads and the second type of functionalized beads has a diameter in a range of between 50 nm and 10 μ

m.

22. The system of claim 21, wherein each of the diameters of the functionalized beads of the first type and the second type is in a range of between 0.5 μ

m and 5 μ

m.

23. The system of claim 21, wherein the diameter of the functionalized beads of the first type is similar to the diameter of the functionalized beads of the second type.

24. The system of claim 21, wherein the diameter of the functionalized beads of the first type is different from the diameter of the functionalized beads of the second type by at least 50%.

25. The system of claim 1, further including a plurality of functionalized beads of at least a third type, functionalized to include at least the second moiety that associates with at least a second analyte under suitable conditions.

26. The system of claim 25, further including a plurality of functionalized beads of a fourth type, functionalized to include the second moiety that associates with the second analyte under suitable conditions.

27. The system of claim 25, wherein the functionalized beads of the first and/or second type further include at least one additional moiety that associates with the second analyte under suitable conditions.

28. The system of claim 27, further including a third moiety that associates with a third analyte under suitable conditions, wherein the functionalized beads of the first type are further functionalized to include the second moiety, and the functionalized beads of the second type are further functionalized to include the third moiety.

3. (canceled)

6. (canceled)

13. (canceled)

29. A method of detecting a complex including an analyte, the method comprising:

(a) contacting a sample in a solution with a population of functionalized beads of a first type, which are magnetic functionalized beads and are functionalized to include a first moiety that can specifically associate with an analyte under appropriate conditions;

(b) contacting the sample solution with a population of functionalized beads of a second type, which are functionalized to include a second moiety that associates with the analyte under suitable conditions, contact resulting in formation of a complex including one of the first type of functionalized bead, the analyte, and one of the second type of functionalized bead; and

(c) detecting the complex including the analyte by detecting magnetic fields produced by the magnetic functionalized bead and by detecting the functionalized bead of the second type associated with the analyte in the complex.

30. The method of claim 29, further including disposing the sample solution including the complex over a substrate that includes at least one optically detected magnetic resonance (ODMR) center formed in the substrate;

exciting electrons within the at least one ODMR center from a ground state to an excited state with incident light;

applying a bias magnetic field on the complex; and

generating a microwave field incident on the at least one ODMR center, the microwave field including frequencies that correspond to ground state transitions in the at least one ODMR center, wherein detecting the complex including the analyte further includes analyzing light emitted by the at least one ODMR center, characteristics of the emitted light being influenced by the microwave field and by the magnetic functionalized bead associated with the analyte in the complex.

31. The method of claim 30, wherein the at least one ODMR center is a nitrogen-vacancy center in a diamond lattice.

32. The method of claim 31, wherein the at least one ODMR center is formed in an upper surface of the substrate, or the at least one ODMR center is a plurality of ODMR centers formed in the upper surface of the substrate.

34. The method of claim 32, wherein analyzing light emitted by the plurality of ODMR centers includes imaging the emitted light.

35. The method of claim 29, further including applying a magnetic field gradient to the sample solution after contacting the sample with the population of functionalized beads of the first type, or after contacting the sample solution with the population of functionalized beads of the second type.

37. The method of claim 29, wherein the population of functionalized beads of the first type and the population of functionalized beads of the second type are added to the sample solution sequentially.

38. The method of claim 29, wherein the functionalized beads of the second type are fluorescent functionalized beads, and the method further includes illuminating the complex with incident light that excites fluorescence within the functionalized beads of the second type and fluorescence imaging of the complex.

39. The method of claim 29, wherein the functionalized beads of the second type are magnetic functionalized beads, including a magnetic property distinguishable from the functionalized beads of the first type.

40. The method of claim 29, further including applying a magnetic field gradient to the sample solution after contacting the sample solution with the functionalized beads of the first and second types.

41. The method of claim 40, further including varying the magnetic field gradient applied to the sample solution.

42. The method of claim 29, further including concentrating the sample solution after contacting the sample solution with the population of functionalized beads of the second type.

43. The method of claim 29, further including agglomerating a plurality of functionalized beads of the first and second types, after contacting the sample solution with the population of functionalized beads of the second type, before detecting the complex.

44. The method of claim 29, further including dehydrating the sample solution after disposing the sample solution over the diamond substrate.

33. (canceled)

36. (canceled)

45-58. -58. (canceled)