Antibodies, kit and method for detecting amyloid beta oligomers
This invention is a selective Aβ oligomer kit and immunoassay method capable of reliably and sensitively detecting Aβ oligomers in a biological sample of a patient. In one embodiment the inventive assay uses a pair of anti-Aβ oligomer antibodies, as capture and detection antibodies, to detect and quantify Aβ oligomers. The method can be used to differentiate Alzheimer's disease (AD) patients from non-AD patients and/or to stratify AD patients according to the severity of their disease.
1. A kit for detecting oligomers of amyloid beta comprising:
(a) a capture antibody that has an affinity for amyloid beta 1-42 oligomers compared to amyloid beta 1-40 monomers in a competitive binding assay of at least 500:1 and has:
a light chain variable region comprising,
(i) a CDR1 having the sequence Arg-Ser-Ser-Gln-Ser-Ile-Val-His-Ser-Xaa 1 -Gly-Xaa 2 -Thr-Tyr-Leu-Glu (SEQ ID NO:1), wherein Xaa 1 is Asn, Ser, Thr, Ala, Asp or Glu and Xaa 2 is Asn, His, Gln, Ser, Thr, Ala, or Asp,
(ii) a CDR2 having the sequence Lys-Ala-Ser-Xaa 1 -Arg-Phe-Ser (SEQ ID NO:2), wherein Xaa 1 is Asn, Gln, Ser, Thr, or Ala, and
(iii) a CDR3 having the sequence Phe-Gln-Gly-Ser-Xaa 1 -Xaa 2 -Xaa 3 -Xaa 4 -Xaa 5 (SEQ ID NO:3), wherein Xaa 1 is Arg, Lys or Tyr, Xaa 2 is Val or Ala, Xaa 3 is Pro or His, Xaa 4 is Ala, Pro, or Val, and Xaa 5 is Ser, Gly, Arg or Phe; and
a heavy chain variable region comprising,
(i) a CDR1 of SEQ ID NO:4,
(ii) a CDR2 of SEQ ID NO:5, and
(iii) a CDR3 of SEQ ID NO:6; or
a light chain variable region comprising,
(i) a CDR1 having the sequence Arg-Ser-Ser-Gln-Ser-Ile-Val-His-Ser-Xaa 1 -Gly-Xaa 2 -Thr-Tyr-Leu-Glu (SEQ ID NO:1), wherein Xaa 1 is Ser, Thr, Ala, Asp or Glu and Xaa 2 is Asn, His, Gln, Ser, Thr, Ala, or Asp,
(ii) a CDR2 having the sequence Lys-Ala-Ser-Xaa 1 -Arg-Phe-Ser (SEQ ID NO:2), wherein Xaa 1 is Asn, Gln, Ser, Thr, or Ala, and
(iii) a CDR3 of SEQ ID NO:16; and
a heavy chain variable region comprising,
(i) a CDR1 of SEQ ID NO:4,
(ii) a CDR2 of SEQ ID NO:5, and
(iii) a CDR3 of SEQ ID NO:6; and
(b) a detection antibody that recognizes an N-terminal linear epitope of amyloid beta 1-42 peptide.
2. The kit of claim 1 , wherein the affinity of the capture antibody for amyloid beta 1-42 oligomers compared to amyloid beta 1-42 monomers in a sandwich ELISA assay is at least 500:1.
3. The kit of claim 1 , wherein the affinity of the capture antibody for amyloid beta 1-42 oligomers compared to amyloid beta 1-42 monomers in a sandwich ELISA assay is at least 1000:1.
4. The kit of claim 1 , wherein the level of detection of amyloid beta 1-42 oligomers by said kit is less than 5 pg/mL.
5. The kit of claim 1 , wherein the level of detection of amyloid beta 1-42 oligomers by said kit is less than 3 pg/mL.
6. The kit of claim 1 , wherein the detection antibody is 6E10, BAM-10, W0-2, 26D6, 2A10, 2B4, 4C2, 4E2, 2H4, 20C2, 2D6, 5F10, 1F4, 1F6, 2E12, 3B3 or 82E1.
7. The kit of claim 1 , wherein the detection antibody further comprises a label.
8. The kit of claim 1 , further comprising a means for concentrating an antibody-antigen complex.
9. An isolated antibody, or antigen binding fragment thereof, that binds amyloid β-derived diffusible ligands, has an affinity for amyloid beta 1-42 oligomers compared to amyloid beta 1-40 monomers in a competitive binding assay of at least 500:1, and has:
(a) a light chain variable region comprising,
(i) a CDR1 having the sequence Arg-Ser-Ser-Gln-Ser-Ile-Val-His-Ser-Xaa 1 -Gly-Xaa 2 -Thr-Tyr-Leu-Glu (SEQ ID NO:1), wherein Xaa 1 is Ser, Thr, Ala, Asp or Glu and Xaa 2 is Asn, His, Gln, Ser, Thr, Ala, or Asp,
(ii) a CDR2 of SEQ ID NO:15, and
(iii) a CDR3 of SEQ ID NO:16; and
(b) a heavy chain variable region comprising,
(i) a CDR1 of SEQ ID NO:4,
(ii) a CDR2 of SEQ ID NO:5, and
(iii) a CDR3 of SEQ ID NO:6.
10. The antibody of claim 9 , wherein the CDR1 of the light chain variable region has the sequence Arg-Ser-Ser-Gln-Ser-Ile-Val-His-Ser-Xaa 1 -Gly-Xaa 2 -Thr-Tyr-Leu-Glu (SEQ ID NO:1), wherein Xaa 1 is Thr, Ala, Asp or Glu and Xaa 2 is Thr.
11. An isolated antibody, or antigen binding fragment thereof, that binds amyloid β-derived diffusible ligands, has an affinity for amyloid beta 1-42 oligomers compared to amyloid beta 1-40 monomers in a competitive binding assay of at least 500:1, and has:
(a) a light chain variable region comprising,
(i) a CDR1 of SEQ ID NO:14,
(ii) a CDR2 having the sequence Lys-Ala-Ser-Xaa 1 -Arg-Phe-Ser (SEQ ID NO:2), wherein Xaa 1 is Gln, Ser, Thr, or Ala, and
(iii) a CDR3 of SEQ ID NO:16; and
(b) a heavy chain variable region comprising,
(i) a CDR1 of SEQ ID NO:4,
(ii) a CDR2 of SEQ ID NO:5, and
(iii) a CDR3 of SEQ ID NO:6.
12. The antibody of claim 11 , wherein the CDR2 of the light chain variable region has the sequence Lys-Ala-Ser-Xaa 1 -Arg-Phe-Ser (SEQ ID NO:2), wherein Xaa 1 is Thr.
13. An isolated antibody, or antigen binding fragment thereof, that binds amyloid β-derived diffusible ligands comprising:
a light chain variable region comprising,
(i) a CDR1 having the sequence Arg-Ser-Ser-Gln-Ser-Ile-Val-His-Ser-Xaa 1 -Gly-Xaa 2 -Thr-Tyr-Leu-Glu (SEQ ID NO:1), wherein Xaa 1 is Ser, Thr, Ala, Asp or Glu and Xaa 2 is Asn, His, Gln, Ser, Thr, Ala, or Asp,
(ii) a CDR2 having the sequence Lys-Ala-Ser-Xaa 1 -Arg-Phe-Ser (SEQ ID NO:2), wherein Xaa 1 is Asn, Gln, Ser, Thr, or Ala, and
(iii) a CDR3 having the sequence Phe-Gln-Gly-Ser-Xaa 1 -Xaa 2 -Xaa 3 -Xaa 4 -Xaa 5 (SEQ ID NO:3), wherein Xaa 1 is Arg, Lys or Tyr, Xaa 2 is Val, Ala or Leu, Xaa 3 is Pro, His or Gly, Xaa 4 is Ala, Pro, or Val, and Xaa 5 is Ser, Gly, Arg or Phe; and
a heavy chain variable region comprising,
(i) a CDR1 of SEQ ID NO:4,
(ii) a CDR2 of SEQ ID NO:5, and
(iii) a CDR3 of SEQ ID NO:6.