A fluorescence-based glucose biosensor using concanavalin A and dextran encapsulated in a poly(ethylene glycol) hydrogel

A fluorescence biosensor is described that is based on a photopolymerized p oly(ethylene glycol) (PEG) hydrogel incorporating fluorescein isothiocyanat e dextran (FITC-dextran) and tetramethylrhodamine isothiocyanate concanaval in A (TRITC-Con A) chemically conjugated into the hydrogel network using an alpha-acryloyl, omega-N-hydroxysuccinimidyl ester of PEG-propionic acid. I n the absence of glucose, TRITC-Con A binds with FITC-dextran, and the FITC fluorescence is quenched through fluorescence resonance energy transfer. C ompetitive glucose binding to TRITC-Con A liberates FITC-dextran, resulting in increased FITC fluorescence proportional to the glucose concentration. In vitro experiments of hydrogel spheres in a solution of 0.1 M phosphate-b uffered saline (pH 7.2) and glucose were conducted for multiple TRITC-Con A /FITC-dextran ratios. Hydrogels were characterized on the basis of the perc ent change in fluorescence intensity when FITC-dextran was liberated by inc reasing glucose concentrations. The optimum fluorescent change between 0 an d 800 mg/dL was obtained with a TRITC-Con A/FITC-dextran mass ratio of 500: 5 mu g/mL PEG. Fluorescent response was linear up to 600 mg/dL, At higher c oncentrations, the response saturated due to the displacement of the majori ty of the FITC-dextran and to concentration quenching by free FITC-dextran, Dynamic fluorescent change upon glucose addition was similar to 10 min for a glucose concentration step change from 0 to 200 mg/dL.