Reusable optical bioassay platform with permeability-controlled hydrogel pads for selective saccharide detection.
A reusable optical bioassay platform using permeability-controlled hydrogel pads for selective saccharide detection has been developed. An optical glucose detection assay based on fluorescence resonance energy transfer (FRET) between dye-labeled dextran and Concanavalin A (ConA) was incorporated into hydrogel pads by entrapment. The hydrogel pads are constructed from hemispherical hydrogel attached onto hydrophobic surfaces of a microtiter plate. The resulted hemispherical hydrogel pads entrapping the sensing biological materials were further surface coated with polyelectrolyte multilayers through a Layer-by-Layer (LbL) self-assembly process to create a permeability-controlled membrane with nanometer thickness. The selective permeable LbL film deposited on the hydrogel surface allows small molecular weight analytes to diffuse into the hydrogel pads while the large molecular weight sensing biological molecules are immobilized. An encapsulation efficiency of 75% for the ConA/Dextran complex within the coated hydrogel pads was achieved and no significant leakage of the complex was observed. Glucose calibration curve with linear range from 0 to 10mM glucose was obtained. Selective permeability of the hydrogel pads has been demonstrated by measurement of saccharides with various molecular weights. The LbL hydrogel pads could selectively detect monosaccharides (glucose, MW=180) and disaccharides (sucrose, MW=342) while polysaccharides (dextran, MW approximately 70kDa) cannot diffuse through the LbL layer and are excluded. LbL hydrogel pads allow regeneration of the FRET system with good signal reproducibility of more than 90% to construct a reusable and reagentless optical bioassay platform.
Cheung
Kwan Yee
KY
Division of Bioengineering, National University of Singapore, Singapore.
Mak
Wing Cheung
WC
Trau
Dieter
D
D001681:Biological Assay / Q000295:instrumentation* / Q000379:methods*; D002241:Carbohydrates / Q000032:analysis*; D003208:Concanavalin A; D003911:Dextrans; D017743:Equipment Reuse; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D005947:Glucose / Q000032:analysis; D020100:Hydrogels / Q000737:chemistry; D007700:Kinetics; D010539:Permeability; D012235:Rhodamines
D002241:Carbohydrates; D003911:Dextrans; D020100:Hydrogels; D012235:Rhodamines; C015219:fluorescein isothiocyanate dextran; D003208:Concanavalin A; D016650:Fluorescein-5-isothiocyanate; D005947:Glucose