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 comprising:
(a) an isolated antibody, or antigen binding fragment thereof, having:
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) an antibody selected from the group consisting of 6E10, BAM-10, W0-2, 26D6, 2A10, 2B4, 4C2, 4E2, 2H4, 20C2, 2D6, 5F10, 1F4, 1F6, 2E12, 3B3 or 82E1 wherein the selected antibody is detectably labeled.
2. A method for detecting oligomers of amyloid beta comprising:
(a) obtaining a biological sample having oligomers of amyloid beta from an animal;
(b) contacting said biological sample with an isolated antibody, or antigen binding fragment thereof, 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 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
under conditions sufficient to form a capture antibody/oligomer of amyloid beta complex;
(c) contacting the complex of step (b) with a detection antibody selected from the group consisting of 6E10, BAM-10, W0-2, 26D6, 2A10, 2B4, 4C2, 4E2, 2H4, 20C2, 2D6, 5F10, 1F4, 1F6, 2E12, 3B3 or 82E1, under conditions sufficient to form capture antibody/oligomer of amyloid beta/detection antibody complex, wherein the detection antibody recognizes an N-terminal linear epitope of amyloid beta 1-42 peptide; and
(d) detecting the complex of step (c).
3. A method of claim 2 , wherein the animal is a human.
4. The method of claim 2 , wherein the biological sample comprises cerebral spinal fluid.
5. The method of claim 2 , wherein said method is capable of detecting less than 5 pg/mL of amyloid beta 1-42 oligomers in the cerebral spinal fluid.
6. The method of claim 2 , wherein said method is capable of detecting less than 3 pg/mL of amyloid beta 1-42 oligomers in the cerebral spinal fluid.
7. The method of claim 2 , wherein the detection antibody further comprises a label.
8. The method of claim 2 , further comprising the step of concentrating the capture antibody/oligomer of amyloid beta/detection antibody complex prior to detecting the complex.