Hydrogel-based protein and oligonucleotide microchips on metal-coated surfaces: enhancement of fluorescence and optimization of immunoassay.
Manufacturing of hydrogel-based microchips on metal-coated substrates significantly enhances fluorescent signals upon binding of labeled target molecules. This observation holds true for both oligonucleotide and protein microchips. When Cy5 is used as fluorophore, this enhancement is 8-10-fold in hemispherical gel elements and 4-5-fold in flattened gel pads, as compared with similar microchips manufactured on uncoated glass slides. The effect also depends on the hydrophobicity of metal-coated substrate and on the presence of a layer of liquid over the gel pads. The extent of enhancement is insensitive to the nature of formed complexes and immobilized probes and remains linear within a wide range of fluorescence intensities. Manufacturing of gel-based protein microarrays on metal-coated substrates improves their sensitivity using the same incubation time for immunoassay. Sandwich immunoassay using these microchips allows shortening the incubation time without loss of sensitivity. Unlike microchips with probes immobilized directly on a surface, for which the plasmon mechanism is considered responsible for metal-enhanced fluorescence, the enhancement effect observed using hydrogel-based microchips on metal-coated substrates might be explained within the framework of geometric optics.
Zubtsova
Zh I
ZhI
Engelhardt Institute of Molecular Biology of Russian Academy of Sciences, Moscow, Russia. zubtsova.z.i@gmail.com
Zubtsov
D A
DA
Savvateeva
E N
EN
Stomakhin
A A
AA
Chechetkin
V R
VR
Zasedatelev
A S
AS
Rubina
A Yu
AY
D000818:Animals; D002138:Calibration; D002232:Carbocyanines / Q000378:metabolism; D002417:Cattle; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000378:metabolism; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000378:metabolism*; D007118:Immunoassay / Q000379:methods*; D008670:Metals / Q000378:metabolism*; D015335:Molecular Probes / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D009841:Oligonucleotides / Q000378:metabolism*; D040081:Protein Array Analysis; D012710:Serum Albumin, Bovine / Q000378:metabolism; D013499:Surface Properties
D002232:Carbocyanines; D005456:Fluorescent Dyes; D008670:Metals; D015335:Molecular Probes; D009841:Oligonucleotides; C085321:cyanine dye 5; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate; D012710:Serum Albumin, Bovine