4.7 Article

Discrimination between protein glycoforms using lectin-functionalised gold nanoparticles as signal enhancers

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NANOSCALE HORIZONS
卷 8, 期 3, 页码 377-382

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ROYAL SOC CHEMISTRY
DOI: 10.1039/d2nh00470d

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Glycoforms of proteins can indicate disease state, making it crucial to develop new tools for detecting protein glycosylation. This study presents a sandwich bio-assay approach using antibodies immobilised on biolayer interferometry sensors to select proteins, and gold nanoparticles functionalised with lectins to identify specific glycoforms. The nanoparticles significantly enhance the signal, allowing the detection of glycoforms at low concentrations without the need for enrichment or manual steps. Proof of concept using prostate specific antigen shows the potential of glycoform analysis in distinguishing cancerous and non-cancerous status.
Glycoforms (and other post-translational modifications) of otherwise identical proteins can indicate pathogenesis/disease state and hence new tools to detect and sense a protein's glycosylation status are essential. Antibody-based assays against specific protein sequences do not typically discriminate between glycoforms. Here we demonstrate a 'sandwich' bio-assay approach, whereby antibodies immobilised onto biolayer interferometry sensors first select proteins, and then the specific glycoform is identified using gold nanoparticles functionalised with lectins which provide signal enhancement. The nanoparticles significantly enhance the signal relative to lectins alone, allowing glycoform specific detection as low as 0.04 mu g mL(-1) (1.4 nM) in buffer, and crucially there is no need for an enrichment step and all steps can be automated. Proof of concept is demonstrated using prostate specific antigen: a biomarker for prostate cancer, where glycoform analysis could distinguish between cancerous and non-cancerous status, rather than only detecting overall protein concentration.

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