Methods of diagnosing ALS
The invention relates to an epitope protection assay for use in diagnosis, prognosis and therapeutic intervention in diseases, for example, involving polypeptide aggregation, such as prion infections. The methods of the invention first block accessible polypeptide target epitope with a blocking agent. After denaturation of the polypeptide, a detecting agent is used to detect protein with target epitope that was inaccessible during contact with the blocking agent. The invention also relates to novel amyotrophic lateral sclerosis-specific epitopes and their uses to make antibodies, and to the novel antibodies and uses thereof.
1. A method of detecting or diagnosing amyotrophic lateral sclerosis in a subject comprising the steps of:
(a) contacting a test sample of said subject with an antibody specific for an ALS-specific epitope, wherein the ALS-specific epitope is selected from the group consisting of:
DLGKGGNEESTKTGNAGS;
(SEQ ID NO: 1)
and
NPLSRKHGGPKDEE
(SEQ ID NO: 2)
to produce an antibody-antigen complex;
(b) measuring the amount of the antibody-antigen complex in the test sample; and
(c) comparing the amount of antibody-antigen complex in the test sample to a control;
wherein a difference in the amount of antibody-antigen complex in the test sample as compared to the control is indicative of amyotrophic lateral sclerosis.
2. The method of claim 1 , wherein the antibody is specific for DLGKGGNEESTKTGNAGS (SEQ ID NO: 1).
3. The method of claim 1 , wherein the antibody is specific for NPLSRKHGGPKDEE (SEQ ID NO:2).
4. The method according to claim 1 , wherein the antibody is monoclonal, polyclonal, chimeric or humanized.
5. The method according to claim 1 , wherein the antibody is an antibody fragment.
6. The method according to claim 5 , wherein the antibody fragment is a Fab, Fab′, F(ab′) 2 , scFv, dsFv, ds-scFv, dimers, minibodies, diabodies, or multimers thereof or bispecific antibody fragments.
7. A kit for diagnosing amyotrophic lateral sclerosis according to the method of claim 1 comprising (1) an antibody specific for an ALS-specific epitope, wherein the ALS-specific epitope comprises an isolated peptide selected from the group consisting of:
DLGKGGNEESTKTGNAGS;
(SEQ ID NO: 1)
and
NPLSRKHGGPKDEE;
(SEQ ID NO: 2)
(2) instructions for the use thereof; and (3) a recombinant peptide standard.
8. The kit of claim 7 wherein the standard is an isolated peptide selected from the group consisting of:
DLGKGGNEESTKTGNAGS;
(SEQ ID NO: 1)
and
NPLSRKHGGPKDEE.
(SEQ ID NO: 2)
9. The kit according to claim 8 , wherein the antibody is specific for DLGKGGNEESTKTGNAGS (SEQ ID NO: 1).
10. The kit according to claim 8 , wherein the antibody is specific for NPLSRKHGGPKDEE (SEQ ID NO:2).
11. A diagnostic agent comprising (1) an antibody specific for an ALS-specific epitope, wherein the ALS-specific epitope comprises an isolated peptide selected from the group consisting of:
DLGKGGNEESTKTGNAGS;
(SEQ ID NO: 1)
and
NPLSRKHGGPKDEE;
(SEQ ID NO: 2)
attached to (2) a label that produces a detectable signal, directly or indirectly wherein the label comprises a radioisotope, a fluorescent compound, a chemiluminescent compound, an enzyme, an imaging agent or a metal ion.
12. The diagnostic agent according to claim 11 , wherein the antibody is specific for DLGKGGNEESTKTGNAGS (SEQ ID NO: 1).
13. The diagnostic agent according to claim 11 , wherein the antibody is specific for NPLSRKHGGPKDEE (SEQ ID NO:2).
14. A kit comprising the diagnostic agent of claim 11 and instructions for the use thereof.