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ConA-based glucose sensing using the long-lifetime azadioxatriangulenium fluorophore

Cummins, Brian and Simpson, Jonathan and Gryczynski, Zygmunt and Sørensen, Thomas Just and Laursen, Bo W. and Graham, Duncan and Birch, David and Coté, Gerard (2014) ConA-based glucose sensing using the long-lifetime azadioxatriangulenium fluorophore. In: Progress in Biomedical Optics and Imaging - Proceedings of SPIE. SPIE Proceedings, 8591 . SPIE, Bellingham. ISBN 9780819498649

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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.