Detection of Amylin-β-amyloid Hetero-Oligomers by Enzyme-Linked Immunosorbent Assay
Amylin is an amyloidogenic neuroendocrine hormone co-synthesized and co-secreted with insulin from the pancreas. It readily crosses the blood–brain barrier and synergistically forms mixed amyloid plaques with β-amyloid (Aβ) in brain parenchyma. Parenchymal amylin-Aβ plaques are found in both sporadi...
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Bio-protocol LLC
2025-02-01
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| author | Noah Leibold Deepak Kotiya Nirmal Verma Florin Despa |
| author_facet | Noah Leibold Deepak Kotiya Nirmal Verma Florin Despa |
| author_sort | Noah Leibold |
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| description | Amylin is an amyloidogenic neuroendocrine hormone co-synthesized and co-secreted with insulin from the pancreas. It readily crosses the blood–brain barrier and synergistically forms mixed amyloid plaques with β-amyloid (Aβ) in brain parenchyma. Parenchymal amylin-Aβ plaques are found in both sporadic and early-onset familial Alzheimer’s disease (AD), yet their (patho)physiological role remains elusive, particularly due to a lack of detection modalities for these mixed plaques. Previously, we developed an enzyme-linked immunosorbent assay (ELISA) capable of detecting amylin-Aβ hetero-oligomers in brain lysate and blood using a polyclonal anti-amylin antibody to capture hetero-oligomers and a monoclonal anti-Aβ mid-domain detection antibody combination. This combination allows for the recognition of distinct amylin epitopes, which remain accessible after amylin-Aβ oligomerization has begun, and precise detection of Aβ epitopes available after oligomer formation. The utility of this assay is evidenced in our previous report, wherein differences in hetero-oligomer content in brain tissue from patients with and without AD and patients with and without diabetes were distinguished. Additionally, using AD model rats, we provided evidence that our assay can be employed for the detection of amylin-Aβ in blood. This assay and protocol are important innovations in the field of AD research because they meet an unmet need to detect mixed amyloid plaques that, if targeted therapeutically, could reduce AD progression and severity. |
| format | Article |
| id | doaj-art-518a6c1ec2404c6b827c85db84753e68 |
| institution | Kabale University |
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| language | English |
| publishDate | 2025-02-01 |
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| spelling | doaj-art-518a6c1ec2404c6b827c85db84753e682025-02-07T08:16:46ZengBio-protocol LLCBio-Protocol2331-83252025-02-0115310.21769/BioProtoc.5179Detection of Amylin-β-amyloid Hetero-Oligomers by Enzyme-Linked Immunosorbent AssayNoah Leibold0Deepak Kotiya1Nirmal Verma2Florin Despa3Department of Pharmacology & Nutritional Sciences, University of Kentucky, Lexington, KY, USAResearch Center on Healthy Metabolism, University of Kentucky, Lexington, KY, USADepartment of Pharmacology & Nutritional Sciences, University of Kentucky, Lexington, KY, USAResearch Center on Healthy Metabolism, University of Kentucky, Lexington, KY, USADepartment of Pharmacology & Nutritional Sciences, University of Kentucky, Lexington, KY, USAResearch Center on Healthy Metabolism, University of Kentucky, Lexington, KY, USADepartment of Pharmacology & Nutritional Sciences, University of Kentucky, Lexington, KY, USAResearch Center on Healthy Metabolism, University of Kentucky, Lexington, KY, USA, Department of Neurology, University of Kentucky, Lexington, KY, USAAmylin is an amyloidogenic neuroendocrine hormone co-synthesized and co-secreted with insulin from the pancreas. It readily crosses the blood–brain barrier and synergistically forms mixed amyloid plaques with β-amyloid (Aβ) in brain parenchyma. Parenchymal amylin-Aβ plaques are found in both sporadic and early-onset familial Alzheimer’s disease (AD), yet their (patho)physiological role remains elusive, particularly due to a lack of detection modalities for these mixed plaques. Previously, we developed an enzyme-linked immunosorbent assay (ELISA) capable of detecting amylin-Aβ hetero-oligomers in brain lysate and blood using a polyclonal anti-amylin antibody to capture hetero-oligomers and a monoclonal anti-Aβ mid-domain detection antibody combination. This combination allows for the recognition of distinct amylin epitopes, which remain accessible after amylin-Aβ oligomerization has begun, and precise detection of Aβ epitopes available after oligomer formation. The utility of this assay is evidenced in our previous report, wherein differences in hetero-oligomer content in brain tissue from patients with and without AD and patients with and without diabetes were distinguished. Additionally, using AD model rats, we provided evidence that our assay can be employed for the detection of amylin-Aβ in blood. This assay and protocol are important innovations in the field of AD research because they meet an unmet need to detect mixed amyloid plaques that, if targeted therapeutically, could reduce AD progression and severity.https://bio-protocol.org/en/bpdetail?id=5179&type=0 |
| spellingShingle | Noah Leibold Deepak Kotiya Nirmal Verma Florin Despa Detection of Amylin-β-amyloid Hetero-Oligomers by Enzyme-Linked Immunosorbent Assay Bio-Protocol |
| title | Detection of Amylin-β-amyloid Hetero-Oligomers by Enzyme-Linked Immunosorbent Assay |
| title_full | Detection of Amylin-β-amyloid Hetero-Oligomers by Enzyme-Linked Immunosorbent Assay |
| title_fullStr | Detection of Amylin-β-amyloid Hetero-Oligomers by Enzyme-Linked Immunosorbent Assay |
| title_full_unstemmed | Detection of Amylin-β-amyloid Hetero-Oligomers by Enzyme-Linked Immunosorbent Assay |
| title_short | Detection of Amylin-β-amyloid Hetero-Oligomers by Enzyme-Linked Immunosorbent Assay |
| title_sort | detection of amylin β amyloid hetero oligomers by enzyme linked immunosorbent assay |
| url | https://bio-protocol.org/en/bpdetail?id=5179&type=0 |
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