28 February 2014 ConA-based glucose sensing using the long-lifetime azadioxatriangulenium fluorophore
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Abstract
Fluorescent glucose sensing technologies have been identified as possible alternatives to current continuous glucose monitoring approaches. We have recently introduced a new, smart fluorescent ligand to overcome the traditional problems of ConA-based glucose sensors. For this assay to be translated into a continuous glucose monitoring device where both components are free in solution, the molecular weight of the smart fluorescent ligand must be increased. We have identified ovalbumin as a naturally-occurring glycoprotein that could serve as the core-component of a 2nd generation smart fluorescent ligand. It has a single asparagine residue that is capable of displaying an N-linked glycan and a similar isoelectric point to ConA. Thus, binding between ConA and ovalbumin can potentially be monovalent and sugar specific. This work is the preliminary implementation of fluorescently-labeled ovalbumin in the ConA-based assay. We conjugate the red-emitting, long-lifetime azadioxatriangulenium (ADOTA+) dye to ovalbumin, as ADOTA have many advantageous properties to track the equilibrium binding of the assay. The ADOTA-labeled ovalbumin is paired with Alexa Fluor 647-labeled ConA to create a Förster Resonance Energy Transfer (FRET) assay that is glucose dependent. The assay responds across the physiologically relevant glucose range (0-500 mg/dL) with increasing intensity from the ADOTA-ovalbumin, showing that the strategy may allow for the translation of the smart fluorescent ligand concept into a continuous glucose monitoring device.
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Brian Cummins, Brian Cummins, Jonathan Simpson, Jonathan Simpson, Zygmunt Gryczynski, Zygmunt Gryczynski, Thomas Just Sørensen, Thomas Just Sørensen, Bo W. Laursen, Bo W. Laursen, Duncan Graham, Duncan Graham, David Birch, David Birch, Gerard Coté, Gerard Coté, } "ConA-based glucose sensing using the long-lifetime azadioxatriangulenium fluorophore", Proc. SPIE 8951, Optical Diagnostics and Sensing XIV: Toward Point-of-Care Diagnostics, 89510A (28 February 2014); doi: 10.1117/12.2039824; https://doi.org/10.1117/12.2039824
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