At a certain angle of incidence, a portion of the light energy through the metal coating couples with the electrons in the metal surface layer, which then move as a result of excitation. Consequently, when there is a small change in the reflective index of the sensing medium (eg, by biomolecule attachment), plasmon cannot be formed. The limit of detection (LOD) in the SPR experiment depends on a number of factors, including the molecular weight, optical property, and binding affinity of target-probe molecules as well as the surface coverage of the probe molecule.
Thus, changes in the intensity of the reflected light are proportional to any change in the refractive index near the metal surface [33]. Due to measurement limitations (fixed wavelength and incidence angle θ), changes in reflectivity (Δ%R) at a single array point can be detected simultaneously. A wide range of applications has been developed for the use of SPR biosensors in the biomedical field.
A detailed review of the use of these DNA analogs as recognition elements in SPR-based sensing can be found elsewhere [54]. This application of the SPR technique has been used to monitor the structural transition in protein-small molecule interactions [12], proteins in different environmental conditions [55, 56] or effects on apoptosis inducers [57]. In SPR imaging, the angle of incidence remains constant, and the binding of biomolecules to the gold surface is measured as the change in reflectance (or reflectance) with the intensity of the incident beam, unlike SPR sensors, which depend on measuring the absorption drop. in SPR angle or SPR wavelength.
Schematic representation of the PSA immunosensor based on PSA detection antibody-modified Au nanoparticles using surface plasmon resonance (SPR).
Immobilization of Biomolecules and Signal Amplification in SPR 1. Immobilization of Biomolecules
Streptavidin-DNA conjugates have been used as molecular linkers in the DNA-directed immobilization of biotinylated antibodies on the chip surface (Figure 5A,B) [97]. Studies of DNA-binding domain-mediated immobilization of proteins on a consensus DNA-modified surface have been reported [99]. Sequence-specific interactions of the c-Myb protein with its specific DNA sequences [ 101 ] and the p53 transcription factor with its consensus DNA binding sequences using SPR techniques have been reported [ 65 ].
The affinity capture strategy is based on modifying the sensor chip to capture specific proteins conjugated with labels. To control the orientation of the immobilized antibody, genetically modified G protein with different affinity tags such as 6ХHis [105], GST [106], elastin [107] and DNA molecules [108] has been developed. Even if the Ni-NTA-6ХHis bond exhibits a weaker binding affinity than the biotin-streptavidin complex, hexahistidine-mediated affinity interaction based on the concept of immobilized metal ion affinity chromatography (IMAC) is one of the most promising methods for orientation. -Controlled immobilization of proteins, as the functional properties of the tagged proteins are only slightly hindered by the small size of the 6ХHis tag.
Instead, some proteins tagged with large affinity tags, such as GST (~25.6 kDa) or MBP (~40.6 kDa) can cause significant steric hindrance, which can lead to in the loss of structural integrity and biological activity of affinity fusion partners. With the advent of recombinant protein techniques, it is possible to express and purify a fusion protein containing the amino acid cysteine with the ease of preparing the cysteine-modified protein. Cysteine-modified G protein-based well-targeted antibody immobilization does not inhibit antibody active sites in response to an antigen; thus, antibody activity on the biosensor surface is unaffected.
Nanoparticle amplification tags are based on the fact that, upon addition of linked Au NPs, a pseudo-mass increase of analyte is induced. The magnetic properties of the beads were used for SEB purification and concentration of real complex matrices before the samples were allowed to flow into their miniature SPR system (Figure 8). One of the properties that make it useful in SPR signal enhancement is its ultrafast electron transport at room temperature.
The charge transfer from the surface of graphene to the surface of the Au thin film makes the sensing surface more sensitive to changes in the sensing medium. The configuration of the proposed graphene-on-gold surface plasmon resonance biosensor based on the generalized N-Layer model, where the gold film is deposited on top of an SF10 glass prism. The graphene-on-gold surface plasmon resonance: prism | Au (50 nm) | graphene (L x 0.34 nm) | detection medium, where L is the number of graphene layers and Z0 = 100 nm is the thickness of the biomolecule layer.
Since SPR-based detection is highly dependent on the mass change of the analytes in the sensing environment, researchers have attempted to achieve the signal. SPR assay for ATP detection amplified by DNA-based hybridization chain reaction (HCR). A) S1 magnetic bead conjugates were prepared by direct immobilization of an amine-modified ATP aptamer linked to an activated carboxylate group on magnetic beads via EDC/NHS chemistry, and then hybridized to its complementary S2 oligonucleotide; (B) Strategy of continuous SPR monitoring of trigger DNA on a DNA chip array.
Recent Advances of SPR Technology
A major advantage of objective-type SPRI is that the optical paths of the sample and imager are fixed during the incidence angle scan. Each of these pixels accordingly produces an SPR curve and the image is framed using the SPR minimum angle information. Following this approach, variations such as inhomogeneity and unwanted interferences from the laser intensity can be significantly removed by using a homogeneous reference surface.
More importantly, the divergence in angles of incidence at different points of the field of view caused by the deviation of the objective lens could also be corrected. First, the use of a high NA, high magnification imaging system could provide diffraction limit resolution (~300 nm) for the imaging optics [133]. Provided that SPR occurs at an angle greater than the critical angle, the immersion objective is chosen with an NA (such as an Olympus 1.65 NA objective) greater than the refractive index of the medium.
Consequently, when the incident light is transmitted to the trailing edge of the objective aperture, it will arrive at the sample surface at an angle greater than the critical angle. Second, compared with the angle-scanning method of conventional SPR measurement, the use of an objective lens could simplify the system design by converting the rotational motion of the incident light on the sample into a linear motion of the stage. By exploiting the advanced new capabilities of SPRI, researchers have successfully applied this high spatial resolution SPR to imaging and detecting single molecules of DNA [135], virus [136], cells [137].
Recent advances in plasmonic imaging techniques using the high-resolution surface plasmon resonance microscopy (SPRM) approach would undoubtedly have far-reaching impacts on the quantitative analysis of intracellular dynamics in living cells, single-molecule analysis, and studies of the biological activities of membrane proteins. such as the discovery of new drugs that target membrane proteins.
Conclusions
Surface plasmon resonance imaging analysis of protein-protein interactions using on-chip expressed capture protein. Use of surface plasmon resonance for studies of protein-protein and protein-phospholipid membrane interactions. Sensing charge state and conformational change in immobilized protein using surface plasmon resonance sensor.
Surface plasmon resonance biosensor for label-free and highly sensitive point mutation detection using extension polymerization reaction. Plasmon resonance imaging as a tool to monitor biomolecular interactions in an array-based format. Surface plasmon resonance imaging-based protein arrays for high-throughput screening of protein-protein interaction inhibitors.
Single-nucleotide polymorphism genotyping by nanoparticle-enhanced surface plasmon resonance imaging measurements of surface binding reactions. High-sensitivity detection of carbohydrate antigen 15-3 using a gold/zinc oxide thin film surface plasmon resonance-based biosensor. Surface plasmon resonance (SPR) sensors for rapid and sensitive detection of cellular response to osmotic stress.
Label-free and time-resolved measurements of cell volume changes by surface plasmon resonance (SPR) spectroscopy. DNA-directed protein immobilization for simultaneous detection of multiple analytes by surface plasmon resonance biosensor. Kinetic analysis of DNA binding by the c-Myb DNA binding domain using surface plasmon resonance.
Isothermal and rapid detection of pathogenic microorganisms using a nano-rolling circle gain-surface plasmon resonance biosensor. A surface plasmon resonance assay coupled to a hybridization chain reaction for enhanced detection of DNA and small molecules. Enzymatically enhanced surface plasmon resonance imaging detection of DNA by exonuclease III digestion of DNA microarrays.
