AMELIORATION OF HETEROPHILE ANTIBODY IMMUNOSENSOR INTERFERENCE

• US 20100248273A1
• Filed: 03/25/2009
• Published: 09/30/2010
• Est. Priority Date: 03/25/2009
• Status: Active Grant

• Current Assignee: Abbott Point Of Care Incorporated (Abbott Laboratories)
• Original Assignee: Abbott Point Of Care Incorporated (Abbott Laboratories)
• Inventors: Omakor, John Emegbero, Campbell, John Lewis Emerson
• Granted Patent: US 8,084,272 B2
• Time in Patent Office: Days
• US Class Current: 435/7.940
• CPC Class Codes: G01N 2333/4712 Muscle proteins, e.g. myosin, actin, protein
• CPC Class Codes: G01N 2333/58 Atrial natriuretic factor complex; Atriopeptin; Atrial natriuretic peptide [ANP]; Brain natriuretic peptide [BNP, proBNP]; Cardionatrin; Cardiodilatin
• CPC Class Codes: G01N 33/5306 Improving reaction conditions, e.g. reduction of non-specific binding, promotion of specific binding
• CPC Class Codes: Y10S 436/824 Immunological separation techniques
• CPC Class Codes: Y10S 436/825 Pretreatment for removal of interfering factors from sample
• CPC Class Codes: Y10T 436/112499 with sample on test slide
• CPC Class Codes: Y10T 436/25125 Digestion or removing interfering materials
• CPC Class Codes: Y10T 436/25375 Liberation or purification of sample or separation of material from a sample [e.g., filtering, centrifuging, etc.]

Abstract

The invention is directed to methods and devices for reducing interference from heterophile antibodies in an analyte immunoassay. In one embodiment, the invention is to a method comprising the steps of (a) amending a biological sample such as a whole blood sample with non-human IgM or fragments thereof by dissolving into said sample a dry reagent to yield a non-human IgM concentration of at least about 20 μg/mL or equivalent fragment concentration; and (b) performing an electrochemical immunoassay on the amended sample to determine the concentration of said analyte in said sample. Preferably, the sample is amended with IgG or fragments thereof in addition to the IgM of fragments thereof.

Claims

1. A method of reducing interference from heterophile antibodies in an analyte immunoassay, comprising:

(a) amending a whole blood sample with non-human IgM or fragments thereof by dissolving into said sample a dry reagent to yield a non-human IgM concentration of at least 20 μ

g/mL or equivalent fragment concentration; and

(b) performing an electrochemical immunoassay on the amended sample to determine the concentration of said analyte in said sample.

2. The method of claim 1, further comprising:

(c) amending the whole blood sample with IgG or fragments thereof.

3. The method of claim 2, wherein the whole blood sample is amended with IgM.

4. The method of claim 2, wherein the whole blood sample is amended with IgM F(ab′

)2 fragments.

5. The method of claim 2, wherein the whole blood sample is amended with IgM Fab fragments.

6. The method of claim 2, wherein the whole blood sample is amended with IgM Fc fragments.

7. The method of claim 2, wherein the analyte is a cardiovascular marker.

8. The method of claim 2, wherein the immunoassay is for an analyte selected from the group, TnI, TnT, CKMB, myoglobin, BNP, NT-proBNP, and proBNP.

9. The method of claim 2, wherein the sample is amended for a predetermined period ranging from about 1 minute to about 30 minutes.

10. The method of claim 2, wherein the dry reagent further comprises a component selected from the group consisting of buffer, salt, surfactant, stabilizing agent, a simple carbohydrate, a complex carbohydrate and combinations thereof.

11. The method of claim 2, wherein the non-human IgG and IgM are murine, caprine or a combination thereof.

12. The method of claim 2, wherein the electrochemical immunoassay is an enzyme-linked sandwich immunoassay.

13. The method of claim 2, wherein the electrochemical immunoassay is performed by an immunosensor.

14. The method of claim 2, wherein the electrochemical immunoassay is performed by an immunosensor and an immuno-reference sensor.

15. The method of claim 2, wherein the dry reagent further comprises an enzyme-labeled antibody to said analyte.

16. The method of claim 2, further comprising:

(d) amending the amended sample with an enzyme-labeled antibody to said analyte by dissolving into said amended sample a second dry reagent comprising said enzyme-labeled antibody, wherein said second dry reagent is separate from the dry reagent that contains said IgM or fragments thereof.

17. The method of claim 2, wherein the electrochemical assay is performed with an immobilized antibody to the analyte on an electrode.

18. The method of claim 2, wherein the amended sample further comprises an enzyme-labeled antibody to said analyte and is contacted with an immobilized antibody to said analyte to form a sandwich of said analyte between said immobilized and labeled antibodies, the method further comprising the steps of washing said sample to a waste chamber and exposing said sandwich to a substrate capable of reacting with said enzyme to form a product capable of electrochemical detection.

19. The method of claim 2, wherein the method is performed at the point of patient care.

20. The method of claim 2, wherein the electrochemical immunoassay is an enzyme-linked immunosorbent assay.

21. The method of claim 2, wherein the immunoassay is performed in a cartridge comprising an immunosensor, a conduit, a sample entry port and a sample holding chamber.

22. The method of claim 21, wherein at least a portion of at least one of said sample entry port, said sample holding chamber, said conduit and said immunosensor are coated with said dry reagent.

23. The method of claim 2, wherein the dry reagent further comprises said non-human IgG.

24. The method of claim 23, wherein the analyte is TnI, and wherein said dry reagent dissolves into the sample to give an IgM concentration of from about 20 to about 200 μ

g/mL and an IgG concentration of from about 50 to about 5000 μ

g/mL.

25. The method of claim 23, wherein the analyte is TnI, and wherein said dry reagent dissolves into the sample to give an IgM concentration of from about 20 to about 60 μ

g/mL and an IgG concentration of from about 500 to about 1000 μ

g/mL.

26. The method of claim 23, wherein the analyte is BNP, and wherein said dry reagent dissolves into the sample to give an IgM concentration of from about 20 to about 200 μ

g/mL and an IgG concentration of from about 50 to about 5000 μ

g/mL.

27. The method of claim 23, wherein the analyte is BNP, and wherein said dry reagent dissolves into the sample to give an IgM concentration of from about 20 to about 60 μ

g/mL and an IgG concentration of from about 500 to about 1000 μ

g/mL.

28. A method of reducing interference from heterophile antibodies in a cardiac troponin I immunoassay, comprising:

(a) amending a whole blood sample with a mixture comprising;

(i) non-human IgG or IgG fragments, and (ii) non-human IgM or IgM fragments, sufficient to substantially sequester any heterophile antibodies in said sample, wherein the non-human IgM concentration in the amended sample is at least about 20 μ

g/mL or equivalent IgM fragment concentration; and

(b) performing an electrochemical immunoassay on the amended sample.

29. A method of reducing interference from heterophile antibodies in a brain natriuretic peptide immunoassay, comprising:

(a) amending a sample with a mixture comprising;

(i) non-human IgG or IgG fragments, and (ii) non-human IgM or IgM fragments, sufficient to substantially sequester any heterophile antibodies in said sample, wherein the non-human IgM concentration in the amended sample is at least about 20 μ

g/mL or equivalent IgM fragment concentration; and

(b) performing an electrochemical immunoassay on the amended sample.

30. A device for performing an immunoassay of an analyte in a blood sample with reduced interference from heterophile antibodies, comprising a housing, an electrochemical immunosensor, a conduit and a sample entry port, wherein said conduit permits a blood sample to pass from the entry port to said immunosensor, and wherein at least one of said housing, said entry port, and said conduit includes a dry reagent comprising non-human IgM or fragments thereof and optionally IgG or fragments thereof, said dry reagent being capable of dissolving into said blood sample to yield an IgM concentration of at least about 20 μ

g/mL or equivalent fragment concentration and substantially sequestering any heterophile antibodies in said sample.

31. The device of claim 30, wherein the dry reagent comprises non-human IgM or fragments thereof and IgG or fragments thereof.

32. The device of claim 31, further comprising a metering system for metering an initial blood sample to form a metered blood sample.

33. The device of claim 31, further comprising an immuno-reference sensor.

34. The device of claim 31, wherein the analyte is a cardiovascular marker.

35. The device of claim 31, wherein the immunoassay is for an analyte selected from the group, TnI, TnT, CKMB, myoglobin, BNP, NT-proBNP, proBNP, beta-HCG, TSH, D-dimer, and PSA.

36. The device of claim 31, wherein the sample is amended for a predetermined period in the range from of about 1 minute to about 30 minutes.

37. The device of claim 31, wherein the device is a single-use cartridge.

38. The device of claim 31, wherein the dry reagent further comprises an enzyme-labeled antibody to the analyte.

39. The device of claim 31, further comprising a second dry reagent comprising an enzyme-labeled antibody to the analyte, wherein the second dry reagent is separate from the dry reagent that comprises the IgM or fragments thereof.

40. The device of claim 31, wherein the dry reagent further comprises a component selected from the group consisting of buffer, salt, surfactant, stabilizing agent, a simple carbohydrate, a complex carbohydrate and combinations thereof.

41. The device of claim 31, wherein the non-human IgG or fragments thereof and IgM or fragments thereof are murine, caprine or a combination thereof.

42. The device of claim 31, wherein the immunosensor performs an electrochemical enzyme-linked sandwich immunoassay.

43. The device of claim 31, wherein the immunosensor comprises an immobilized antibody to the analyte on an electrode.

44. The device of claim 31, wherein the analyte is TnI, and wherein said dry reagent dissolves into the sample to give an IgM concentration of from about 20 to about 200 μ

g/mL and an IgG concentration of from about 50 to about 5000 μ

g/mL.

45. The device of claim 31, wherein the analyte is TnI, and wherein said dry reagent dissolves into the sample to give an IgM concentration of from about 20 to about 60 μ

g/mL and an IgG concentration of from about 500 to about 1000 μ

g/mL.

46. The device of claim 31, wherein the analyte is BNP, and wherein said dry reagent dissolves into the sample to give an IgM concentration of from about 20 to about 200 μ

g/mL and an IgG concentration of from about 50 to about 5000 μ

g/mL.

47. The device of claim 31, wherein the analyte is BNP, and wherein said dry reagent dissolves into the sample to give an IgM concentration of from about 20 to about 60 μ

g/mL and an IgG concentration of from about 500 to about 1000 μ

g/mL.

48. The device of claim 31, further comprising a wash fluid capable of washing said sample to a waste chamber.

49. The device of claim 31, further comprising a wash fluid comprising a substrate capable of reacting at said immunosensor to form a product capable of electrochemical detection.

50. A method of reducing interference from heterophile antibodies in an analyte immunoassay comprising:

(a) amending a biological sample with IgM or fragments thereof and optionally IgG or fragments thereof to yield a non-human IgM concentration of at least about 20 μ

g/mL or equivalent fragment concentration; and

(b) performing an immunoassay on the amended sample to determine the concentration of said analyte in said sample.

51. The method of claim 50, further comprising:

(c) amending the biological sample with IgG or fragments thereof.

52. The method of claim 51, wherein the biological sample is selected from the group consisting of whole blood, serum, plasma, urine and diluted forms thereof.

53. The method of claim 51, wherein the biological sample is amended by dissolving into said sample a dry reagent comprising IgM.

54. The method of claim 51, wherein the biological sample is amended by dissolving into said sample a dry reagent comprising IgM fragments.

55. The method of claim 51, wherein the biological sample is amended by adding a solution comprising IgM or fragments thereof.

56. The method of claim 51, wherein the immunoassay method is selected from the group consisting of electrochemical, amperometric, potentiometric, absorbance, fluorescence and luminescence.

57. A method of reducing heterophile antibody interference in an analyte immunoassay device, comprising:

(a) adding IgM or fragments thereof and IgG or fragments thereof to a biological sample in an amount sufficient to substantially sequester any heterophile antibodies in said sample and forming an amended sample, wherein the IgM or fragments thereof and the IgG or fragments thereof are added at a weight ratio greater than 0.004; and

(b) performing an electrochemical immunoassay on said amended sample to determine the concentration of the analyte in said amended sample.

58. The method of claim 57, wherein said weight ratio is greater than 0.02.

59. The method of claim 57, wherein said weight ratio is greater than 0.05.

60. The method of claim 57, wherein step (a) comprises adding IgM to said biological sample.

61. The method of claim 60, wherein after the adding step, the IgM is present in said sample in a concentration of at least 20 μ

g/mL.

62. The method of claim 57, wherein step (a) comprises adding IgM fragments to said biological sample.

63. The method of claim 62, wherein after the adding step, the IgM fragments are present in said sample in a concentration equivalent to an IgM concentration of at least about 20 μ

g/mL.

64. The method of claim 57, wherein the adding comprises dissolving the IgM or the fragments thereof into said sample from a dry reagent coating contained in the immunoassay device.

65. The method of claim 57, wherein the adding comprises dissolving the IgM or the fragments thereof into said sample from a dry reagent coating contained on a sample collection device.

66. The method of claim 57, wherein the adding comprises mixing said sample with a liquid comprising said IgM or said fragments thereof to form an amended mixture, the method further comprising the step of introducing the amended mixture into the immunoassay device.

67. A device for performing an immunoassay of an analyte in a blood sample with reduced interference from heterophile antibodies, comprising a housing, an electrochemical immunosensor, a conduit and a sample entry port, wherein said conduit permits a blood sample to pass from the entry port to said immunosensor, and wherein at least one of said housing, said entry port, and said conduit includes a dry reagent comprising non-human IgM or fragments thereof and IgG or fragments thereof at a weight ratio greater than 0.004.

68. The device of claim 67, wherein the weight ratio is greater than 0.02.

69. The device of claim 67, wherein the weight ratio is greater than 0.05.