Microfluidic systems integrated with two-dimensional surface plasmon resonance phase imaging systems for microarray immunoassay

This study reports a microfluidic chip integrated with an arrayed immunoassay for surface plasmon resonance (SPR) phase imaging of specific bio-samples. The SPR phase imaging system uses a surface-sensitive optical technique to detect two-dimensional (2D) spatial phase variation caused by rabbit immunoglobulin G (IgG) adsorbed on an anti-rabbit IgG film. The microfluidic chip was fabricated by using micro-electro-mechanical-systems (MEMS) technology on glass and polydimethylsiloxane (PDMS) substrates to facilitate well-controlled and reproducible sample delivery and detection. Since SPR detection is very sensitive to temperature variation, a micromachine-based temperature control module comprising micro-heaters and temperature sensors was used to maintain a uniform temperature distribution inside the arrayed detection area with a variation of less than 0.3 degrees C. A self-assembled monolayer (SAM) technique was used to pattern the surface chemistry on a gold layer to immobilize anti-rabbit IgG on the modified substrates. The microfluidic chip is capable of transporting a precise amount of IgG solution by using micropumps/valves to the arrayed detection area such that highly sensitive, highly specific bio-sensing can be achieved. The developed microfluidic chips, which employed SPR phase imaging for immunoassay analysis, could successfully detect the interaction of anti-rabbit IgG and IgG. The interactions between immobilized anti-rabbit IgG and IgG with various concentrations have been measured. The detection limit is experimentally found to be 1 x 10(-4)mg/ml (0.67 nM). The specificity of the arrayed immunoassay was also explored. Experimental data show that only the rabbit IgG can be detected and the porcine IgG cannot be adsorbed. The developed microfluidic system is promising for various applications including medical diagnostics, microarray detection and observing protein-protein interactions.     

Real-time protein biosensor arrays based on surface plasmon resonance differential phase imaging

This paper reports the application of differential phase surface plasmon resonance (SPR) imaging in two-dimensional (2D) protein biosensor arrays. Our phase imaging approach offers a distinct advantage over the conventional angular SPR technique in terms of utilization efficiency of optical sensor elements in the imaging device. In the angular approach, each biosensor site in the biosensor array requires a linear array of optical detector elements to locate the SPR angular dip. The maximum biosensor density that a two-dimensional imaging device can offer is a one-dimensional SPR biosensor array. On the other hand, the phase-sensitive SPR approach captures data in the time domain instead of the spatial domain. It is possible that each pixel in the captured interferogram represents one sensor site, thus offering high-density two-dimensional biosensor arrays. In addition, our differential phase approach improves detection resolution through removing common-mode disturbances. Experimental results demonstrate a system resolution of 8.8 x 10(-7)RIU (refractive index unit). Real-time monitoring of bovine serum albumin (BSA)/anti-BSA binding interactions at various concentration levels was achieved using a biosensor array. The detection limit was 0.77 microg/ml. The reported two-dimensional SPR biosensor array offers a real-time and non-labeling detection tool for high-throughput protein array analysis. It may find promising applications in protein therapeutics, drug screening and clinical diagnostics.     

Phase-polarisation contrast for surface plasmon resonance biosensors

A technique of phase-polarisation contrast (PPC) for the enhancement of the contrast of a surface plasmon resonance (SPR) intensity profile is proposed and experimentally realised. The technique exploits the peculiarities of light phase and polarisation behaviour under SPR. It applies to non-optimum SPR coupling conditions and enables one to lower the resonant minimum of reflected intensity nearly to zero, and hence to increase substantially the ratio of the intensity from the resonance to that at the minimum. We observed the contrast enhancement by more than one order of magnitude when we applied the PPC scheme. The PPC can be efficiently employed in commercial SPR sensors, as it significantly reduces restrictions on allowable parameters of SPR-supporting metal films and biomolecular layers immobilised on them, facilitates SPR observation, and increases the accuracy of SPR shift measurements.     

Phase-polarisation contrast for surface plasmon resonance biosensors

A technique of phase-polarisation contrast (PPC) for the enhancement of the contrast of a surface plasmon resonance (SPR) intensity profile is proposed and experimentally realised. The technique exploits the peculiarities of light phase and polarisation behaviour under SPR. It applies to non-optimum SPR coupling conditions and enables one to lower the resonant minimum of reflected intensity nearly to zero, and hence to increase substantially the ratio of the intensity from the resonance to that at the minimum. We observed the contrast enhancement by more than one order of magnitude when we applied the PPC scheme. The PPC can be efficiently employed in commercial SPR sensors, as it significantly reduces restrictions on allowable parameters of SPR-supporting metal films and biomolecular layers immobilised on them, facilitates SPR observation, and increases the accuracy of SPR shift measurements.     

Surface Plasmon Resonance Based Fiber Optic Ammonia Sensor Utilizing Bromocresol Purple

In this paper, a surface plasmon resonance (SPR) based fiber optic ammonia gas sensor has been designed and fabricated using bromocresol purple (BCP) as sensing element. The sensor works under wavelength modulation scheme. The detection of ammonia gas has been carried out at room temperature. Three different kinds of film coating configurations, namely silver + BCP, gold + BCP, and silver + silicon + BCP on the unclad portion of the fiber have been used for studying the role of each layer. Further, to optimize the performance of the sensor, the films of varying thicknesses were coated using thermal evaporation technique. Experiments have been performed for the ammonia concentrations ranging from 0 to 150 ppm around the probe. To record the SPR spectrum, light from a polychromatic source is launched in the fiber and the spectrum is recorded at the other end of the fiber. The spectrum has a peak at lower wavelength while a dip at the higher wavelength. The dip corresponds to SPR while the peak appears to be due to fluorescence properties of the dye. It has been observed that as the ammonia gas comes in contact of the BCP layer, it changes the refractive index of the BCP dye which, in turn, changes the resonance wavelength. Further, the change in refractive index increases as the concentration of ammonia gas increases up to certain concentration of ammonia after that it saturates. Silicon layer has been shown as a protection layer for silver and gold from oxidation and acts as a tuner of wavelength. The proposed ammonia sensor has small response as well as recovery time.     

A new approach for the detection of DNA sequences in amplified nucleic acids by a surface plasmon resonance biosensor

In this paper, a simple and useful approach for DNA sensing based on surface plasmon resonance (SPR) transduction is reported. A new DNA sample pre-treatment has been optimised to allow fast and simple detection of hybridisation reaction between a target sequence in solution and a probe immobilised on the sensing surface. This pre-treatment consisted in a denaturation procedure of double stranded DNA containing the target sequence and was based on an high temperature treatment (95 degrees C, 5 min) followed by a 1 min incubation with small oligonucleotides. The oligonucleotides are designed to prevent the re-hybridising of the denatured strands, while enabling the target sequence to bind the immobilised probe. The important parameters of the procedure, i.e. incubation time, length and concentration of the oligonucleotides, have been studied in detail. The optimised DNA denaturation procedure has been successfully applied to the detection of amplified DNA with a commercially available SPR biosensor (Biacore X). DNA samples extracted from plant and human blood were tested after amplification by polymerase chain reaction (PCR).     

Sub-attomole oligonucleotide and p53 cDNA determinations via a high-resolution surface plasmon resonance combined with oligonucleotide-capped gold nanoparticle signal amplification

Oligonucleotide (ODN)-capped gold nanoparticles (Au-NPs) were used in a sandwich assay of ODN or polynucleotide by a flow injection surface plasmon resonance (SPR). A carboxylated dextran film was immobilized onto the SPR sensor surface to eliminate nonspecific adsorption of ODN-capped Au-NPs. The tandem use of signal amplification via the adlayer of the ODN-capped Au-NPs and the differential signal detection by the bicell detector on the SPR resulted in a remarkable DNA detection level. A 39-mer target at a quantity as low as 2.1 x 10(-20)mol, corresponding to 1.38 fM in a 15 microl solution, can be measured. To our knowledge, both the concentration and quantity detection levels are the lowest among all the gene analyses conducted with SPR to this point. The method is shown to be reproducible (relative standard deviation values <16%) and to possess high sequence specificity. It is also demonstrated to be viable for sequence-specific p53 cDNA analysis. The successful elimination of nonspecific adsorption of, and the signal amplification by, ODN-capped Au-NPs renders the SPR attractive for cases where the DNA concentration is extremely low and the sample availability is severely limited.     

Analysis of mono- and oligosaccharides by multiwavelength surface plasmon resonance (SPR) spectroscopy.

Surface plasmon resonance (SPR) spectra of different saccharides were collected using a home-made multiwavelength SPR apparatus. Pentoses, hexoses, disaccharides and a trisaccharide were distinguished from one another according to their SPR spectra collected at the same concentration. The spectra were also used for the quantitation of sugars by exploring the linear relationship between resonance wavelength and solute concentration. The dynamic linear ranges for the determination of glucose, sucrose and raffinose are 0.01-0.2, 0.005-0.1 and 0.0025-0.1 mol/L, respectively. The SPR spectrum of a mixture of two components was investigated. While the experiments have not been carried out, the implications from this work are that the technique would be applicable to mixtures containing more than two components.     

Biosensing based on surface plasmon resonance and living cells

We propose the combination of surface plasmon resonance (SPR) with living cells as a biosensing method. Our detection scheme is based on the premise that cellular activity induced by external agents is often associated with changes in cellular morphology, which in turn should lead to a variation of the effective refractive index at the interface between the cell membrane and the metal layer. We monitored surface plasmon resonance signals originating from a gold surface coated with cells on a custom apparatus after injection of various agents known to influence cellular activity and morphology. Specifically, we evaluated three types of stimulation: response to an endotoxin (lipopolysaccharides), a chemical toxin (sodium azide) and a physiological agonist (thrombin). A comparison with phase contrast microscopy reveals that SPR signal variations are associated with the induction of cell death for lipopolysaccharides treatment and a contraction of the cell body for sodium azide. Thrombin-induced cellular response shows a rapid decrease of the measured laser reflectance over 5min followed by a return to the original value. For this treatment, phase contrast micrographs relate the first phase of the SPR variation to cell contraction and increase of the intercellular gaps, whereas the recovery phase can be associated with a spreading of the cell on the sensing surface. Hence, the SPR signal is very consistent with the cellular response normally observed for these treatments. This confirms the validity of the biosensing method, which could be applied to a large variety of cellular responses involving shape remodeling induced by external agents.     

Integrated-optic biosensor by electro-optically modulated surface plasmon resonance

We present a new integrated-optic surface plasmon resonance (SPR) biosensor based on electro-optical modulation. The SPR characteristics for the analyte concentration detection can be electro-optically modulated by applying the voltage on the electrodes of the biosensor fabricated on lithium niobate, which is an excellent electro-optic material. Two measurement methods, electro-optically modulated SPR spectral measurement and electro-optically modulated SPR intensity measurement, are demonstrated and their measurands are the SPR wavelength and the output optical intensity, respectively. Human serum albumin is coated on the gold film surface of the proposed biosensor to detect the concentration of beta-blocker, which is a remedy for heart disease. As the applied voltage increases such that the effective index of guided mode rises, the SPR wavelength shifts toward the long wavelength side and the output optical intensity at the wavelength of 632.8 nm diminishes. The linear regression slope of the relation between the measurand and the applied voltage is dependent on the analyte concentration and can be used to determine the concentration variation. Experimental results measured by the electro-optically modulated SPR methods are compared with those measured by the conventional spectral and intensity methods, and the effects of waveguide width on the biosensor performance are discussed.     

Bayesian Estimation of the Active Concentration and Affinity Constants Using Surface Plasmon Resonance Technology

Surface plasmon resonance (SPR) has previously been employed to measure the active concentration of analyte in addition to the kinetic rate constants in molecular binding reactions. Those approaches, however, have a few restrictions. In this work, a Bayesian approach is developed to determine both active concentration and affinity constants using SPR technology. With the appropriate prior probabilities on the parameters and a derived likelihood function, a Markov Chain Monte Carlo (MCMC) algorithm is applied to compute the posterior probability densities of both the active concentration and kinetic rate constants based on the collected SPR data. Compared with previous approaches, ours exploits information from the duration of the process in its entirety, including both association and dissociation phases, under partial mass transport conditions; do not depend on calibration data; multiple injections of analyte at varying flow rates are not necessary. Finally the method is validated by analyzing both simulated and experimental datasets. A software package implementing our approach is developed with a user-friendly interface and made freely available.     

D-Shaped Photonic Crystal Fiber–Based Surface Plasmon Resonance Biosensors with Spatially Distributed Bimetallic Layers

This paper deals with the development and analysis of D-Shaped photonic crystal fiber (PCF) biosensors using surface plasmon resonance (SPR). A thin metal layer is deposited on the outer flat surface of the PCF that behaves as the plasmonic material. Analyte is filled in the outermost peripheral region of metal layer. Finite element method (FEM) with perfectly matched layer (PML) is applied to analyze the proposed sensors. Mode analysis is performed on the proposed structures to evaluate various parameters of SPR-based PCF sensors. Three D-shaped PCF structures have been proposed with silver (Ag), gold (Au) and two-half layers of both (Ag-Au) on its flat surface. The first two structures are analyzed to the range of wavelength where the SPR will occur to facilitate understanding of the third structure. It is observed that the structures with one metal have only one sensitive plasmonic peak whereas the structure with two metal layers has two sensitive plasmonic peaks, making it suitable candidate for two-molecule sensing present in a sample analyte. Good sensitivities and resolutions are achieved for both plasmonic peaks.

     

Metallic Nanowire Array–Polymer Hybrid Film for Surface Plasmon Resonance Sensitivity Enhancement and Spectral Range Enlargement

In this paper, the coupling interaction is investigated between a metallic nanowire array and a metal film under the Kretschmann condition. The plasmonic multilayer is composed of a metallic nanowire array embedded in a polymer layer positioned above a metal film, exploiting the classical surface plasmon resonance (SPR) configuration. We analyze the influence of various structural parameters of the metallic nanowire array on the SPR spectrum of thin metal film. The results show that the coupling interactions of nanowires with the metal film can greatly affect SPR resonance wavelength and increase SPR sensitivity. The coupling strength of metallic nanowire array and metal film also impacts resonance wavelength, which can be used to adjust SPR range but have little effect on its sensitivity. The results are confirmed using a dipole coupling resonance model of metallic nanowire. We demonstrated that this nanostructured hybrid structure can be used for high sensitivity SPR monitoring in a large spectral range, which is important for advanced SPR measurement including fiber-optic SPR sensing technology.     

Wide dynamic range phase-sensitive surface plasmon resonance biosensor based on measuring the modulation harmonics

In this study, a novel phase-sensitive surface plasmon resonance (SPR) setup, based on temporal modulation of a pumping beam by a photoelastic modulator, and subsequent extraction of phase information at the second and the third harmonics of the modulation frequency, has been developed to study biomolecular interactions on SPR-supporting gold. We demonstrated that the design setup provides ultra-high phase sensitivity, together with a wide dynamic range of measurements. In particular, the proposed scheme was used to study real-time interaction of biotin-protein and streptavidin-BSA complexes. We have found that the proposed technique has a detection limit as high as 2.89 x 10(-7) in terms of refractive index units (RIU). In terms of biosensing performance, a detection sensitivity of 1.3 nM from the streptavidin-maleimide/thiolated BSA complex binding reaction has also been demonstrated.     

Surface plasmon resonance sensors in cell biology: basics and application

Optical sensors based on the excitation of surface plasmons, referred to as surface plasmon resonance (SPR) sensors, have become a central analytical tool for characterizing and quantifying a wide variety of macromolecular interactions, like receptor–ligand contacts. Besides this classical field of application, in the last 15 years, the development of SPR sensors aiming for the detection and analysis of ligand/cell or host/pathogen interactions, cell/cell contacts, and cellular reactions gained considerable momentum. The number of publications reporting about applications of SPR sensors implementing vital prokaryotic or eukaryotic cells as biorecognition elements for medical diagnostics, environmental monitoring, or biological safety is steadily growing. This review gives a short introduction to the technique of surface plasmon resonance and the parameters that are important for its application in the field of vital cell sensors. Furthermore, the publications concerning the application of such sensors in the analysis of cellular interactions and cellular reactions to extra- and intracellular stimuli are summarized.     

Temperature-Regulated Surface Plasmon Resonance Imaging System for Bioaffinity Sensing

We describe a temperature-regulated surface plasmon resonance (SPR) imaging biosensor in this article. The sample temperature can be regulated for specific requirements of the bioaffinity sensing, and stabilized to suppress the measurement noise caused by temperature fluctuations. The water thermo optic coefficient is measured to test the temperature regulation performance. The protein interaction is monitored to demonstrate the feasibility of this system for real-time biomolecular interaction analysis. This temperature-regulated SPR imaging biosensor can be readily implemented by adding the common water path and peristaltic pump to the conventional SPR imaging system, which may provide an economical and convenient scheme to improve the analysis accuracy and quality of bioaffinity sensing using SPR sensing platform.     

Surface plasmon resonance: a useful technique for cell biologists to characterize biomolecular interactions

Surface plasmon resonance (SPR) is a powerful technique for monitoring the affinity and selectivity of biomolecular interactions. SPR allows for analysis of association and dissociation rate constants and modeling of biomolecular interaction kinetics, as well as for equilibrium binding analysis and ligand specificity studies. SPR has received much use and improved precision in classifying protein–protein interactions, as well as in studying small-molecule ligand binding to receptors; however, lipid–protein interactions have been underserved in this regard. With the field of lipids perhaps the next frontier in cellular research, SPR is a highly advantageous technique for cell biologists, as newly identified proteins that associate with cellular membranes can be screened rapidly and robustly for lipid specificity and membrane affinity. This technical perspective discusses the conditions needed to achieve success with lipid–protein interactions and highlights the unique lipid–protein interaction mechanisms that have been elucidated using SPR. It is intended to provide the reader a framework for quantitative and confident conclusions from SPR analysis of lipid–protein interactions.     

Line Shape Analysis and Extended Instrumental Operation of Surface Plasmon Resonance Sensors

Line shape effects of the surface plasmon resonance (SPR) at the gold-water interface have been exploited, towards extending instrumental performance in the angular interrogation mode. Comprising a micro-fluidic set-up both, SPR-line broadening and associated signal symmetry/asymmetry features have been monitored in real time for selected adsorption systems and are compared with the simultaneously taken resonance angle that represents the commonly recorded SPR feature. Four angular broadening contributions have been identified: an intrinsic, temperature independent fraction, a temperature dependent part that dominates at ambient temperature, a surface morphology/roughness contribution and an adsorption related fraction. The latter, and substantially smaller contribution, is crucially affected by surface conditions, as the transition from hydrophobic to a hydrophilic surface state. Anionic sub-monolayer adsorption has been clearly resolved, along with detection of a pronounced calorimetric effect that originates from an exothermic surface reaction upon exposure of the protein covered gold surface to a strong chemical oxidant.     

Measurements of the cross-bridge attachment/detachment process within intact sarcomeres by surface plasmon resonance.

We have developed a surface plasmon resonance (SPR) system to monitor the cross-bridge attachment/detachment process within intact sarcomeres from mouse heart muscle. SPR occurs when laser light energy is transferred to surface plasmons that are resonantly excited in a metal (gold) film. This resonance manifests itself as a minimum in the reflection of the incident laser light and occurs at a characteristic angle. The angle of the SPR occurrence depends on the dielectric permittivity of the sample medium adjacent to the gold film. Purified sarcomeric preparations are immobilized onto the gold film in the presence of a relaxing solution. Replacement of the relaxing solution with increasing Ca(2+) concentration solution activates the cross-bridge interaction and produces an increase in the SPR angle. These results imply that the interaction of myosin heads with actin within an intact sarcomere changes the dielectric permittivity of the sarcomeric structure. In addition, we further verify that SPR measurements can detect the changes in the population of the attached cross-bridges with altered concentrations of phosphate, 2,3-butanedione monoxime, or adenosine triphosphate at a fixed calcium concentration, which have been shown to reduce the force and increase the cross-bridge population in attached state. Thus, our data provide the first evidence that the SPR technique allows the monitoring of the cross-bridge attachment/detachment process within intact sarcomeres.     

Nine surface plasmon resonance assays for specific protein quantitation during cell culture and process development

Quantitation of protein is essential during pharmaceutical development, and a variety of methods and technologies for determination of total and specific protein concentration are available. Here we describe the development of a streamlined assay platform for specific quantitation assays using surface plasmon resonance (SPR) technology. A total of nine different assays were developed using similar conditions, of which eight assays were for quantitation of different human blood plasma proteins (IgG, IgG1–4 subclasses, IgA, transferrin, and albumin) from a chromatography-based IgG plasma process. Lastly, an assay for monitoring the concentration of a recombinant monoclonal antibody during 13 days of CHO cell culturing was developed. Assay performances were compared with enzyme-linked immunosorbent assay (ELISA), nephelometry, ARCHITECT, and Cobas c501. SPR assays were shown to have higher sensitivity than analysis using nephelometry, ARCHITECT, and Cobas and to have significantly lower analysis and hands-on time compared with ELISA. Furthermore, the SPR assays were robust enough to be used for up to 12 days, allowing specific protein concentration measurement of a sample to be completed at line within 10 min. Using the same platform with only few varied parameters between different assays has saved time in the lab as well as for evaluation and presentation of results.