Fiber optic SPR biosensing of DNA hybridization and DNA-protein interactions.
In this paper we present a fiber optic surface plasmon resonance (SPR) sensor as a reusable, cost-effective and label free biosensor for measuring DNA hybridization and DNA-protein interactions. This is the first paper that combines the concept of a fiber-based SPR system with DNA aptamer bioreceptors. The fibers were sputtered with a 50nm gold layer which was then covered with a protein repulsive self-assembled monolayer of mixed polyethylene glycol (PEG). Streptavidin was attached to the PEG's carboxyl groups to serve as a versatile binding element for biotinylated ssDNA. The ssDNA coated SPR fibers were first evaluated as a nucleic acid biosensor through a DNA-DNA hybridization assay for a random 37-mer ssDNA. This single stranded DNA showed a 15 nucleotides overlap with the receptor ssDNA on the SPR fiber. A linear calibration curve was observed in 0.5-5 microM range. A negative control test did not reveal any significant non-specific binding, and the biosensor was easily regenerated. In a second assay the fiber optic SPR biosensor was functionalized with ssDNA aptamers against human immunoglobulin E. Limits of detection (2nM) and quantification (6nM) in the low nanomolar range were observed. The presented biosensor was not only useful for DNA and protein quantification purposes, but also to reveal the binding kinetics occurring at the sensor surface. The dissociation constant between aptamer and hIgE was equal to 30.9+/-2.9nM. The observed kinetics fully comply with most data from the literature and were also confirmed by own control measurements.
BIOSYST-MeBioS, Katholieke Universiteit Leuven, Willem de Croylaan 42, B-3001 Leuven, Belgium. jeroen.pollet@biw.kuleuven.be
Jeroen
J
Pollet
Filip
F
Delport
Kris P F
KP
Janssen
Karolien
K
Jans
Guido
G
Maes
Helge
H
Pfeiffer
Martine
M
Wevers
Jeroen
J
Lammertyn
D015374:Biosensing Techniques / Q000295:instrumentation*; D004247:DNA / Q000032:analysis* / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000032:analysis / Q000737:chemistry*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D005336:Fiber Optic Technology / Q000295:instrumentation*; D017403:In Situ Hybridization / Q000295:instrumentation*; D025941:Protein Interaction Mapping / Q000295:instrumentation*; D020349:Surface Plasmon Resonance / Q000295:instrumentation*
D004268:DNA-Binding Proteins; D004247:DNA