Surface Plasmon Resonance Imaging Measurements of DNA Hybridization Adsorption and Streptavidin/DNA Multilayer Formation at Chemically Modified Gold Surfaces

Abstract

A combination of scanning and imaging surface plasmon resonance (SPR) experiments is used to characterize DNA hybridization adsorption at gold surfaces and the subsequent immobilization of streptavidin. Single-stranded oligonucleotides are immobilized at gold surfaces, and the hybridization of biotinylated complements from solution is monitored with SPR. The subsequent attachment of streptavidin to the biotinylated complements provides a method of enhancing the SPR imaging signal produced as a result of the hybridization and leads to a 4-fold improvement in the hybridization detection limit of the SPR imaging apparatus. In situ scanning SPR experiments are used to measure a 60 ± 20% hybridization efficiency between immobilized single-stranded DNA and biotinylated complements. From the information provided by both the in situ imaging and scanning SPR experiments, an absolute surface coverage of immobilized single-stranded DNA is estimated to be ∼3 × 1012 molecules/cm2. The SPR signal resulting from hybridization onto immobilized probes is further amplified by the formation of streptavidin/DNA multilayers which grow by a combination of DNA hybridization and biotin−streptavidin binding. DNA/DNA multilayers without streptavidin are used as an additional method of amplifying the SPR signal.

Discovery and verification of functional single nucleotide polymorphisms in regulatory genomic regions: Current and developing technologies

“…This technique has been utilized to measure many kinds of biomolecular interactions including antibody-epitope recognition (i.e. protein-protein interaction) [53], DNA: DNA or DNA:RNA oligomerization [54][55][56][57] and DNA-protein interactions [58][59][60][61]. DNA microarray technology has been combined with SPR imaging analysis to create a medium to high-throughput methodology for quantifying DNA-protein interactions [62] in which a recently modified DNA array generation process has enhanced detection of interactions with double-stranded sequences of similar makeup (i.e., SNPs in binding regions) [63].…”

Development of Nanostructured Plasmonic Substrates for Enhanced Optical Biosensing

“…Its distinguishing feature is the use of CCD camera where the SPR signals are captured as images after passing through a narrow band interference filter [83]. While SPR imaging scheme has an important advantage of simplicity of structure with no moving parts, the intensity-based method has difficulties in achieving significant detection accuracy because of insufficient sensitivity.…”

Recognizing Hybrid/Mixed G‐quadruplex in Human Telomeres by Using a Cyanine Dye Supramolecule with Confocal Laser Scanning Microscopy

“…32 A variation of resonance wavelength shift (∆λ) of the slides would be observed as the modification reaction progresses. Once ∆λ reached a certain value and kept unchanged, indicating the surfaces had reached the maximum coverage, 33 the assembling was stopped and the Au films were rinsed with ethanol and ultrapure water to thoroughly remove the unbound MUA. The Au films then were immersed in an aqueous solution of 75 mmol•L -1 EDC and 15 mmol• L -1 NHS for 30 min, 34 and thoroughly rinsed with ultrapure water and ethanol before use.…”