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Title: Developing a Europium-based optical biosensor for detection of protein biomarkers
Author: Al-Enezi, Eiman Ali
ISNI:       0000 0004 9351 7980
Awarding Body: University of Leeds
Current Institution: University of Leeds
Date of Award: 2020
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Protein biomarker measurement has been well established using ELISA which offers good sensitivity and specificity but remains slow and expensive. Biosensors offer the prospect of reagent-less, processing-free measurements at the patient's bedside while recent developments in synthetic binding proteins have facilitated biosensor fabrication. The trivalent Europium ion (Eu3+) is a lanthanide that is attractive for use in optical biosensors due to its bright fluorescence when chelated by organic ligands. The aim of this project was to develop a platform for biosensing based on chelated Eu3+ against a range of proteins biomarkers for muscle injury/ heart attack, ischemic stroke, inflammation and colorectal cancer. Affimers against the biomarkers (human myoglobin, glial fibrillary acidic protein (GFAP), C-reactive protein (CRP) and carcinoembryonic antigen (CEA)) were used as targeting bioreceptors. The Affimers were purified and checked for selective binding using ELISA and immunoprecipitation assays. The Eu+3 was then chelated by pyromellitic dianhydride (PMDA) modified Affimers to form Eu+3 -Affimer complexes. The fluorescence characteristics of the complexes against the protein biomarkers were measured in diluted human serum at λex = 395 nm and λem = 590 nm and 615 nm. The Eu3+- Affimer complexes allowed sensitive detection of myoglobin, GFAP, CRP and CEA proteins as low as 100 fM in 1 % (v/v) human serum in 2 to 3 minutes. The Eu3+ complex was far more sensitive against CRP and CEA proteins as compared to GFAP and human myoglobin, and sandwich assays, using an Affimer against the biomarker plus a polyclonal antibody, suggested that the mass of the biomarker may have a substantial effect on quenching of the Eu3+ complex luminescence. These data are the first to demonstrate that the Affimer based Eu3+ complexes can function as nano-biosensors with potential analytical and diagnostic applications.
Supervisor: Millner, Paul ; Jose, Gin ; Saha, Sikha Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available