Novel Technologies for Highly Sensitive Protein Quantification in Blood Plasma and Extracellular Vesicles

Momenbeitollahi, Nikan
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University of Guelph

Proteins and extracellular vesicles (EVs) in biofluids can be used as crucial biomarkers to diagnose diseases early, which can be lifesaving. This thesis describes the development of sensitive and simple technologies to detect and quantify proteins and EVs in human blood plasma. Firstly, a method using a 3D nitrocellulose membrane and gold nanoparticles (AuNPs) was developed, which detected fluorescently-labeled IgG protein spiked in human plasma with a limit of detection of 29 ng/mL, three orders of magnitude lower than that of conventional 2D assays and one order of magnitude lower than 3D membrane assays without using AuNPs. Secondly, a novel EV dot blotting method was developed using a 3D nitrocellulose membrane, with AuNPs added for further signal enhancement. This method showed that a limit of detection (LOD) of 1242 EVs/sample for CD9 protein, 281 EVs/sample for CD24, and 538 EVs/sample for EpCAM were achieved, up to four orders of magnitude lower than conventional ELISA. Both these platforms offer highly sensitive, inexpensive, and simple methods of protein detection, which is crucial for early disease diagnostics.

protein detection, sensitivity enhancement, fluorescence enhancement, extracellular vesicle surface protein quantification, improving limit of detection
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