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Cited by 3Monoclonal and Polyclonal Antibodies ResearchAdvanced Glycation End Products researchDiabetes Management and Research

Towards detection of structurally-diverse glycated epitopes in native proteins: Single-chain antibody directed to non-A1c epitope in human haemoglobin

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Gaetani, Miss Lucia, Pinto, Isabel Campos, Li, Meng, O’Connor, Peter, Giorgi-Coll, Susan, Tyreman, Matthew, Rumary, Katherine L., Schouten, James A. +2 more·2024·Molecular Immunology
DOI 10.1016/j.molimm.2023.12.008
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Abstract

Over 500 million people worldwide are affected by diabetes mellitus, a chronic disease that leads to high blood glucose levels and causes severe side effects. The predominant biological marker for diagnosis of diabetes is glycated haemoglobin (GHb). In human blood the predominant reducing sugar, glucose, irreversibly conjugates onto accessible amine groups within Hb. Most methods for diagnosis and monitoring of diabetes selectively detect N-terminal glycation at Val-1 on the β-globin chain, but not glycation at other sites. Detection of other glycated epitopes of GHb has the potential to provide new information on the extent, duration and timing of elevated glucose, facilitating personalised diagnosis and intelligent diabetic control. In this work, a new anti-GHb Fab antibody (Fab-1) specific for haemoglobin A1c (HbA1c) with nanomolar affinity was discovered via epitope-directed immunisation and phage display. A single chain variable fragment (scFv) antibody derived from Fab-1 retained affinity and specificity for HbA1c, and affinity was enhanced tenfold upon addition of an enhanced green fluorescent protein tag. Both the scFv and Fab-1 recognised an epitope within HbA1c that was distinct from β-Val-1, and our data suggest that this epitope may include glycation at Lys-66 in the β-globin chain. To our knowledge, this is the first report of an scFv/Fab anti-glycated epitope antibody that recognises a non-A1c epitope in GHb, and confirms that fructosamine attached to different, discrete glycation sites within the same protein can be resolved from one another by immunoassay.

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  • Immunoassay-based methods depend on antibody specificity for HbA1c, making them more vulnerable to epitope changes, masking, or variant interference.

    “By contrast, immunoassay-based methods depend on antibody specificity for HbA1c, making them more vulnerable to epitope changes, masking, or variant interference (40).”

    From Comparison of point‑of‑care HbA1c devices with laboratory HPLC methods: A systematic review and meta‑analysis (2026) · cites this paper as [40]

    The Introduction and Discussion and conclusions (page numbers unavailable) describe HbA1c tests as recognizing a particular glycated epitope, while §3.2 evaluates antibody specificity in an ELISA. Section 3.6 shows that chemical alteration or denaturation of HbA1c prevented recognition by the antibody studied. The paper does not establish that immunoassays are more vulnerable than other methods, or specifically document masking or haemoglobin-variant interference. (LLM, conf=0.84)

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Authors
Gaetani, M. L., Pinto, I. C., Li, M., O’Connor, P., Giorgi-Coll, S., Tyreman, M., Rumary, K. L., Schouten, J. A., Davis, P., & Dixon, A. M.
Affiliations
University of Warwick (GB)
Corresponding author
Dixon, Ann M.
ORCID
Venue
Molecular Immunology
Publisher
Elsevier BV
Published
2024-02
Volume
166
Pages
16-28
ISSN
0161-5890 (ISSN-L)
DOI
10.1016/j.molimm.2023.12.008
OpenAlex
W4390575897
Cited by
3
References
46
Citations by year
2026: 1 · 2025: 1 · 2024: 1
FWCI
0.77
Type
journal article
Language
en
Open access
Licence
Locations
  • Molecular Immunology (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Warwick Research Archive Portal (University of Warwick) (acceptedVersion)
Funders
  • University of Warwick
  • Medical Research Council (MR/R015910/1)
Topics
  • Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Monoclonal and Polyclonal Antibodies Research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Clinical Biochemistry › Advanced Glycation End Products research
  • Health Sciences › Medicine › Endocrinology, Diabetes and Metabolism › Diabetes Management and Research
Keywords
HbA1c, scFv, Fab antibody, fructosamine, glycation sites, diabetes diagnosis, phage display, immunoassay
MeSH terms
Epitopes, Diabetes Mellitus, Glucose, Glycated Hemoglobin, Humans, Sodium Oxybate, beta-Globins, Single-Chain Antibodies

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