Method for diagnosing Lyme disease using a cellular immunological test
The present invention relates to a method for diagnosing Lyme disease in a subject, the method comprising the steps of: (a) obtaining a sample from said subject, (b) contacting said sample with a source of Borrelia antigens and (c) determining the expression level of a pro-inflammatory cytokine in said sample at the end of step (b).
1. A method for diagnosing Lyme disease in a subject, the method comprising the steps of:
(a) contacting a sample obtained from said subject with a source of Borellia antigen from a species of Borellia selected from the group consisting of B. burgdorferi, B. garinii, B. afzelii and combinations thereof, wherein said sample comprises peripheral blood mononuclear cells (PBMC);
(b) determining a protein expression level of IL-1β in said sample at the end of step (a); and
(c) diagnosing Lyme disease in said subject when a detectable expression level or an increase of the expression level of IL-1β is determined in step (b).
2. A method according to claim 1 , wherein the protein expression level of IL-1β is determined by directly quantifying an amount of said IL-1β in said sample using a molecule which specifically binds to IL-1β.
3. A method according to claim 1 , wherein said source of Borrelia antigen is a whole Borrelia cell.
4. A method according to claim 1 , wherein the sample is a fluid obtained from the subject.
5. A method according to claim 4 , wherein the fluid is blood.
6. A method according to claim 1 , wherein in (a) if several species of Borrelia are used as a source of Borrelia antigens, the sample of step (a) is divided into several sub-samples, each sub-sample being contacted with a source of antigens from one species of Borrelia.
7. A method according to claim 2 , wherein the molecule which specifically binds IL-1β is an antibody.
8. A device method according to claim 2 , wherein step (b) is performed using a lateral flow test strip which comprises the molecule which specifically binds to IL-1β.
9. A method according to claim 3 , wherein the Borrelia cell is heat-inactivated or formalin-fixated.
10. A method according to claim 1 , wherein the protein expression level of IL-1β is determined by directly quantifying an amount of said IL-1β in said sample using an immunoassay.
11. A method according to claim 10 , wherein said immunoassay is selected from the group consisting of western blotting, ELISA and FACS using latex beads.
12. A method according to claim 10 , wherein said immunoassay is a sandwich immunoassay.
13. A method according to claim 1 , wherein in (c) Lyme disease is diagnosed in said subject when a detectable expression level of the protein expression level of IL-1β is determined in step (b).
14. A method according to claim 13 , wherein the detectable expression level of the protein expression level of IL-1β in (b) is compared to a reference value.
15. A method according to claim 14 , wherein said reference value is determined in a control sample.
16. A method according to claim 13 , wherein the detectable expression level of the protein expression level of IL-1β in (b) is greater than 40 pg/mL.
17. A method according to claim 1 , wherein in (c) Lyme disease is diagnosed in said subject when an increase of the protein expression level of IL-1β is determined in step (b).
18. A method according to claim 17 , wherein the increase of the protein expression level of IL-1β is determined by comparison with a protein expression level of IL-1β in a control sample.
19. A method according to claim 1 , wherein the sample is diluted blood.
20. A method according to claim 1 , wherein the sample comprises peripheral blood mononuclear cells (PBMC) that have been isolated by centrifugation of diluted blood obtained from the subject.
21. A method according to claim 1 , wherein in (a) the contacting has a duration of at least 4 hours.
22. A method according to claim 21 , wherein in (a) the contacting has a duration of 4-96 hours.
23. A method according to claim 3 , wherein in (a) at least 10 4 whole Borrelia cells are contacted with said sample.
24. A method according to claim 1 , wherein the species of Borellia is selected from the group consisting of strain ATCC 35210 of B. burgdorferi , strain ATCC 51383 of B. garinii , strain ATCC 51567 of B. afzelii , and combinations thereof.
25. A method according to claim 3 , wherein the Borrelia cell is heat-killed.
26. A method according to claim 1 , wherein said source of Borrelia antigen is part of a Borrelia cell.
27. A method according to claim 26 , wherein said part of a Borrelia cell is a protein, a digest of the protein and/or a fragment thereof.
28. A method according to claim 26 , wherein said part of Borrelia cell is a lysate, sonicate or fixate of a Borrelia cell.
29. A method according to claim 1 , wherein the detectable expression level or the increase of the expression level of IL-1β is determined by comparison to a reference value.
30. A method according to claim 1 , wherein the detectable expression level or the increase of the expression level of IL-1β is determined by comparison to a control sample.
31. A method for diagnosing Lyme disease in a subject, the method comprising the steps of:
(a) contacting a sample obtained from said subject with a source of Borellia antigen from a species of Borellia selected from the group consisting of B. burgdorferi, B. garinii, B. afzelii and combinations thereof, wherein said sample comprises peripheral blood mononuclear cells (PBMC);
(b) determining protein expression of IL-1β in said sample at the end of step (a); and
(c) diagnosing Lyme disease in said subject when detectable expression of IL-1β is determined in step (b).