Methods for detecting antibodies

• US 10,359,432 B2
• Filed: 12/28/2015
• Issued: 07/23/2019
• Est. Priority Date: 03/27/2008
• Status: Active Grant

• Current Assignee: Regents of the University of California (University of California)
• Original Assignee: Regents of the University of California (University of California)
• Inventors: Kipps, Thomas J., Messmer, Bradley T., Sanchez, Ana B., Kummel, Andrew C., Ruidiaz, Manuel
• Primary Examiner(s): Counts, Gary
• Application Number:
  US14/981,715
• Publication Number:
  US 20160146829A1
• Time in Patent Office: 1,303 Days
• Field of Search: None
• CPC Class Codes: G01N 33/537 with separation of immune complex from unbound antigen or antibody
• CPC Class Codes: G01N 33/54326 Magnetic particles
• CPC Class Codes: G01N 33/577 involving monoclonal antibodies binding reaction mechanisms characterised by the use of monoclonal antibodies; monoclonal antibodies per se are classified with their corresponding antigens; G01N33/53 - G01N33/576 take precedence
• CPC Class Codes: G01N 33/582 with fluorescent label
• CPC Class Codes: G01N 33/6854 Immunoglobulins
• CPC Class Codes: G01N 33/94 involving narcotics or drugs or pharmaceuticals, neurotransmitters or associated receptors

Abstract

Complexes comprising a therapeutic monoclonal antibody and a peptide are provided. In some embodiments, the complexes may comprise a therapeutic monoclonal antibody which is not complexed to an epitope of a target protein and a peptide complexed to the therapeutic monoclonal antibody. In some embodiments, the peptide may have a length of about 5-40 amino acids and may comprise a mimetope recognized by the therapeutic monoclonal antibody, wherein the mimetope comprises a linear sequence of amino acids which is different than a linear sequence of amino acids in the epitope of the target protein.

Claims

1. A complex comprising a therapeutic monoclonal antibody and a peptide, said complex comprising:

a) a therapeutic monoclonal antibody which is not complexed to an epitope of a target protein; and

b) a peptide comprising an amino acid sequence selected from the group consisting of SEQ ID NOs;

1-3, wherein the peptide is complexed to said therapeutic monoclonal antibody, said peptide having a length of about 5-15 amino acids and comprising a mimetope recognized by the therapeutic monoclonal antibody,

wherein the mimetope comprises a linear sequence of amino acids which is different than a linear sequence of amino acids in the epitope of the target protein, wherein the therapeutic monoclonal antibody is alemtuzumab obtained from the circulation of a subject undergoing alemtuzumab treatment, and wherein the peptide is attached directly on a solid support.

2. The complex of claim 1, wherein the mimetope binds to the antigen binding site of the therapeutic monoclonal antibody.

3. The complex of claim 1, wherein the therapeutic monoclonal antibody which is not complexed to an epitope of a target protein is present in a biological sample obtained from a subject.

4. The complex of claim 2, wherein the uncomplexed therapeutic monoclonal antibody is at concentrations from about 50 ng/ml to about 500 ng/ml.

5. The complex of claim 1, wherein the mimetope is identified from a phage-displayed phage library.

6. The complex of claim 1, further comprising a detectable label on the therapeutic monoclonal antibody or the peptide.

7. The complex of claim 6, wherein the detectable label is detectable by a method selected from the group consisting of:

Western blot analysis, flow cytometry, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), competition immunoassay, dual antibody sandwich assay, chemiluminescent assay, bioluminescent assay, fluorescent assay, and agglutination assay.

8. The complex of claim 3, wherein the biological sample is a fluid.

9. The complex of claim 1, wherein the solid support is a bead.

10. The complex of claim 9, wherein the bead is magnetic.

11. The complex of claim 9, wherein the bead comprises magnetic nanoparticles.

12. The complex of claim 11, wherein the magnetic nanoparticles comprise iron oxide (FeO).

13. The complex of claim 11, wherein the bead has a particle size of about 1 μ

m to 50 μ

m.

14. The complex of claim 13, wherein the bead has a particle size of about 10 μ

m.

15. The complex of claim 11, wherein the bead further comprises a fluorophore.

16. The complex of claim 3, wherein the subject is a human subject.

17. The complex of claim 3, wherein the complex comprises a therapeutic monoclonal antibody which is present at concentrations below 500 ng/ml in the biological sample.