Peptides and methods for the detection of lyme disease antibodies
The invention provides compositions (e.g., peptide compositions) useful for the detection of antibodies that bind to Borrelia antigens. The peptide compositions comprise polypeptide sequences comprising variants in the IR6 domain of the Borrelia VlsE protein. The invention also provides devices, methods, and kits comprising such peptide compositions and useful for the detection of antibodies that bind to Borrelia antigens and the diagnosis of Lyme disease.
1. A mixture of isolated peptides comprising three or more different isolated peptides, wherein at least one of said isolated peptides comprises the sequence of SEQ ID NO: 3, SEQ ID NO: 6, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 18, SEQ ID NO: 27, SEQ ID NO: 30, SEQ ID NO: 71, SEQ ID NO: 75, or SEQ ID NO: 100.
2. The mixture of claim 1 , wherein each isolated peptide is conjugated to a ligand.
3. The mixture of claim 1 , wherein one or more of the isolated peptides is biotinylated.
4. The mixture of claim 1 , wherein one or more of the isolated peptides is conjugated to streptavidin or bovine serum albumin optionally via an additional terminal amino acid linking residue.
5. The mixture of claim 1 , wherein each isolated peptide is immobilized to a solid support.
6. A method for detecting in a sample an antibody to an epitope of a Borrelia antigen, the method comprising: contacting a sample with the mixture of isolated peptides of claim 1 and a labeling reagent; and detecting formation of complexes comprising an antibody, the labeling reagent and peptides in the mixture, wherein formation of said complexes is indicative of an antibody to an epitope of a Borrelia antigen being present in said sample.
7. The method of claim 6 , wherein said Borrelia antigen is from a Borrelia burgdorferi, Borrelia afzelli , or Borrelia garinii species.
8. The method of claim 6 , wherein each isolated peptide in the mixture is immobilized to a solid support.
9. The method of claim 8 , wherein said solid support is a bead, a flow path in a lateral flow assay device, a well in a microtiter plate, or a flow path in a rotor.
10. The method of claim 6 , wherein said detecting step comprises (i) performing an ELISA assay, (ii) running a lateral flow assay, (iii) performing an agglutination assay, (iv) performing a Western blot, a slot blot, or dot blot, or (v) running the sample through an analytical rotor.
11. The method of claim 6 , wherein said sample is from a human, canine, or equine subject.
12. The method of claim 6 , wherein said sample is a blood, serum, cerebral spinal fluid, urine, or saliva sample.
13. A method for diagnosing Lyme disease in a subject, the method comprising: contacting a sample from the subject with the mixture of isolated peptides of claim 1 and a labeling reagent; and detecting formation of complexes comprising an antibody labeling reagent and said peptides in the mixture, wherein formation of the complexes is indicative of the subject having Lyme disease.
14. A kit comprising the mixture of isolated peptides of claim 1 and a labeling reagent capable of binding to an antibody that recognizes an epitope of said one or more isolated peptides.
15. The kit of claim 14 , wherein each isolated peptide in the mixture is attached to a solid support.
16. The kit of claim 14 , wherein each isolated peptide in the mixture is attached to a bead, a tube, a well, a lateral flow assay device, or an analytical rotor.
17. The kit of claim 14 , wherein the labeling reagent is
(i) an anti-human or anti-canine IgG antibody conjugated to a detectable label or
(ii) protein A and/or protein G conjugated to a detectable label.
18. The kit of claim 17 , wherein the detectable label is colloidal gold nanoparticles.
19. The mixture of claim 4 , wherein the terminal amino acid linking residue is cysteine or lysine.
20. The mixture of claim 1 , wherein each isolated peptide is attached to or immobilized on a bead, a flow path in a lateral flow immunoassay device, a well in a microtiter plate, or a flow path in a rotor.
21. The mixture of claim 1 , wherein one or more of the isolated peptides is conjugated to a colloidal nanoparticle.
22. The mixture of claim 21 , wherein the colloidal nanoparticle is a colloidal gold nanoparticle.
23. The mixture of claim 1 , wherein each isolated peptide comprises an additional N-terminal peptide sequence that is a native VlsE sequence or non-VlsE Borrelia antigen.
24. The mixture of claim 23 , wherein the additional N-terminal peptide sequence is a sequence of n 1 -n 2 -S-P-n 5 -n 6 -P (SEQ ID NO: 149) or a fragment thereof, wherein n 1 is an amino acid selected from the group consisting of A and V, n 2 is an amino acid selected from the group consisting of E and D, n 5 is an amino acid selected from the group consisting of K and R, and n 6 is an amino acid selected from the group consisting of K and R.
25. The mixture of claim 1 or claim 23 , wherein each isolated peptide comprises an additional C-terminal peptide sequence that is a native VlsE sequence or non-VlsE Borrelia antigen.
26. The mixture of claim 25 , wherein the additional C-terminal peptide sequence is a sequence of V-c 2 -E-G-c 5 -Q-Q-E-G-A-Q-Q-P-S-C (SEQ ID NO: 150) or a fragment thereof, wherein c 2 is an amino acid selected from the group consisting of Q and R, and c 5 is an amino acid selected from the group consisting of V and A.
27. The mixture of claim 25 , wherein the additional C-terminal peptide sequence is a sequence of A-V-c 3 -E-G-c 6 -Q-Q-E-G-A-Q-Q-P-S (SEQ ID NO: 151) or a fragment thereof, wherein c 3 is an amino acid selected from the group consisting of Q and R, and c 6 is an amino acid selected from the group consisting of V and A.
28. A mixture of isolated peptides comprising all possible peptides defined by L-K-K-D-D-N-I-A-A-A-X 11 -V-L-R-G-X 16 -X 17 -K-D-G-X 21 -F-A-X 24 -X 25 (SEQ ID NO: 1) wherein X 11 is an amino acid selected from the group consisting of V and L, X 16 is an amino acid selected from the group consisting of L and I, X 17 is an amino acid selected from the group consisting of A and V, X 21 is an amino acid selected from the group consisting of R, D and N, X 24 is an amino acid selected from the group consisting of I, W, and Y, and X 25 is an amino acid selected from the group consisting of K and R.