Development of instrumentation to allow the detection of microorganisms using light scattering in combination with surface plasmon resonance

This paper describes work carried out to develop a biosensor which allows two separate detection principles to operate simultaneously at the same surface. A prototype device was constructed that provided Kretschmann-configuration surface plasmon resonance (SPR) measurement of refractive index (RI) changes using an 820 nm LED light source, whilst a 635 nm diode laser was used to produce light scattering signals from bacterial spores. Both effects occurred at a gold-coated surface. The RI changes were measured conventionally from the side of the gold layer nearer to the light sources. The scattered light was imaged from the opposite face which was in contact with the aqueous sample. Specific detection of bacterial spores through the light scattering mode using antibody capture was investigated. The flow dynamics and interactions with the surface of individual spores were observed. A comparison with SPR for detection using the same antibody/antigen pair was made. Spore suspensions that were readily detectable by light scattering at 10(7) ml(-1) did not provide significant responses by SPR. The potential for future developments is discussed.     

A new surface plasmon resonance sensor for high-throughput screening applications

We report a new high-throughput surface plasmon resonance (SPR) sensor based on combination of SPR imaging with polarization contrast and a spatially patterned multilayer SPR structure. We demonstrate that this approach offers numerous advantageous features including high-contrast SPR images suitable for automated computer analysis, minimum crosstalk between neighboring sensing channels and inherent compensation for light level fluctuations. Applications of a laboratory prototype of the high-throughput SPR sensor with 108 sensing channels for refractometry and biosensing are described. In refractometric experiments, the noise-limited refractive index resolution of the system has been established to be 3 x 10(-6) refractive index unit (RIU). Experimental data on detection of human choriogonadotropin (hCG) suggest that in conjunction with monoclonal antibodies against hCG, the reported SPR imaging sensor is capable of detecting hCG at concentrations lower than 500 ng/ml.     

Bacteria detection using disposable optical leaky waveguide sensors

Novel disposable absorbing material clad leaky waveguide sensor devices (LWD) have been developed for the detection of pathogenic particles such as bacteria. These chips are tailored to give the maximum extension of the evanescent field at the sensor surface in order to place the entire volume of the bacteria captured by immobilized antibodies on the chip surface within this field. This in turn increases the interaction of the light with the bacteria's bulk volume. Disposable LWD chips were fabricated at room temperature and without the use of expensive fabrication equipment. These LWDs have been characterised by detecting refractive index (RI) changes, scattering and fluorescence from bacterial spores at the sensor surface when illuminated at the coupling angle. The detection limit of Bacillus subtilis var. niger (BG) bacterial spores was 10(4) spores/ml and the illumination intensity of the spores was found to be three times greater than the illumination intensity generated using the surface plasmon resonance (SPR).     

Optimization of Surface Plasmon Resonance Biosensor with Ag/Au Multilayer Structure and Fiber-Optic Miniaturization

In this paper, we report a novel wavelength interrogation-based surface plasmon resonance (SPR) system, in which a film of three Ag layers and three Au layers are alternately deposited on a Kretschmann configuration as sensing element. This multilayer film shows higher sensitivity for refractive index (RI) measurement by comparing with single Au layer structure, which is consistent with its theoretical calculation. A sensitivity range of 2056–5893 nm/RIU can be achieved, which is comparable to RI sensitivities of other wavelength-modulated SPR sensors. Compared with Ag film, this Ag/Au multilayer arrangement offers anti-oxidant protection. This SPR biosensor based on a cost-effective Ag/Au multilayer structure is applicable to the real-time detection of specific interactions and dissociation of low protein concentrations. To extend the application of this highly-sensitive metal film device, we integrated this concept on an optical fiber. The range of RI sensitivities with Ag/Au multilayer was 1847–3309 nm/RIU. This miniaturized Ag/Au multilayer-based fiber optic sensor has a broad application in chemical and biological sensing.     

dc-Sheet resistance as sensitive monitoring tool of protein immobilization on thin metal films

The suitability of high resolution, in situ dc-sheet resistance monitoring (SRM) as a simplified and reliable sensing technique towards detection and tracking of protein immobilization has been explored. Non-specific adsorption of bovine serum albumin (BSA) onto a very thin gold film, acting as the sensing resistor, has been employed as a model system. For comparison, the novel sensing method was combined with surface plasmon resonance (SPR) spectroscopy, using the same flow cell and sensing surface. Two different, well known adsorption states, involving a composite layer of irreversibly and reversibly bound BSA, were clearly resolved by both methods. Clearly structured, pronounced and fully reproducible film resistance modulations have been resolved in the associated SRM data. The transition from reversibly bound BSA to the diluted protein phase is associated with an unusually large decrease in the dc-sheet resistance. The observed resistance modulation magnitude for an adsorbed BSA monolayer corresponds to approximately 1%, and up to 100 mOmega at a 10 Omega sensing resistor. The sheet resistance of irreversibly bound BSA was determined to 0.24 kOmega/cm2, and the associated specific resistivity estimated to 1-2x10(4) Omega cm.     

Multi-analyte surface plasmon resonance biosensing

Surface plasmon resonance (SPR) biosensors are affinity sensing devices exploiting a special mode of electromagnetic field-surface plasmon-polariton-to detect the binding of analyte molecules from a liquid sample to biomolecular recognition elements immobilized on the surface of the sensor. In this paper, we review advances of SPR biosensor technology towards detection systems for the simultaneous detection of multiple analytes (multi-analyte detection). In addition, we report application of a recently developed multichannel SPR sensor based on spectroscopy of surface plasmons and wavelength division multiplexing of sensing channels to multi-analyte detection.     

Rapid method for detection of Salmonella in milk by surface plasmon resonance (SPR)

The Plasmonic surface plasmon resonance (SPR) device was used to develop a rapid, simple and specific immunoassay for detection of Salmonella in milk. Rapid detection of Salmonella contamination is a major challenge for the food industry. Salmonella contamination is well known in all foods including pasteurised milk. The SPR assay was developed as a sandwich model using a polyclonal antibody against Salmonella as capture and detection antibody. Milk spiked with Salmonella typhimurium cells, killed by thimerosal (1%, w/w) treatment was used. Using the Plasmonic SPR assay it was possible to detect S. typhimurium down to a concentration of 1.25 x 10(5) cells ml(-1) in both milk and buffer system. The results obtained are comparable with existing, approved rapid Salmonella detection techniques. No negative effects on the sensitivity of the assay are encountered due to the milk matrix. Hence, no sample preparation or clean-up steps are required. The sample volume requirement for the assay is only 10 microl. Using the assay S. typhimurium was detected in milk within 1h, whereas the cultural techniques require 3-4 days for presumptive positive isolates and further time for confirmation. The rapid tests require at least 24h for the results. The Plasmonic SPR device operates on the Kretschmann configuration and is a cuvette-based system with the advantage of having eight channels on one single SPR chip.     

New trends in instrumental design for surface plasmon resonance-based biosensors

Surface plasmon resonance (SPR)-based biosensing is one of the most advanced label free, real time detection technologies. Numerous research groups with divergent scientific backgrounds have investigated the application of SPR biosensors and studied the fundamental aspects of surface plasmon polaritons that led to new, related instrumentation. As a result, this field continues to be at the forefront of evolving sensing technology. This review emphasizes the new developments in the field of SPR-related instrumentation including optical platforms, chips design, nanoscale approach and new materials. The current tendencies in SPR-based biosensing are identified and the future direction of SPR biosensor technology is broadly discussed.     

A miniature fiber optic surface plasmon resonance sensor for fast detection of Staphylococcal enterotoxin B

A fiber optic surface plasmon resonance (SPR) biosensor for detection of Staphylococcal enterotoxin B (SEB) is reported. The sensor is based on spectral interrogation of surface plasmons in a miniature sensing element based on a side-polished single-mode optical fiber with a thin metal overlayer. For specific detection of SEB, the SPR sensor is functionalized with a covalently crosslinked double-layer of antibodies against SEB. The SPR biosensor is demonstrated to be able to detect ng/ml concentrations of SEB in less than 10 min.     

A polycarbonate based surface plasmon resonance sensing cartridge for high sensitivity HBV loop-mediated isothermal amplification

In this study, we report a simple, low-cost surface plasmon resonance (SPR)-sensing cartridge based on a loop-mediated isothermal amplification (LAMP) method for the on-site detection of the hepatitis B virus (HBV). For LAMP detection, a SPR based LAMP sensing system (SPRLAMP) was constructed, including a novel SPRLAMP sensing cartridge integrating a polymethyl methacrylate (PMMA) micro-reactor with a polycarbonate (PC)-based prism coated with a 50 nm Au film. First, we found that the change of refractive index of the bulk solution was approximately 0.0011 refractive index (RI) units after LAMP reaction. The PC-based prism's linearity and thermal responses were compared to those of a traditional glass prism to show that a PC-based prism can be used for SPR measurement. Finally, the HBV template mixed in the 10 μl LAMP solution could be detected by SPRLAMP system in 17 min even at the detection-limited concentration of 2 fg/ml. We also analyzed the correlation coefficients between the initial concentrations of HBV DNA templates and the system response (ΔRU) at varying amplification times to establish an optimal amplification time endpoint of 25 min (R(2)=0.98). In conclusion, the LAMP reaction could be detected with the SPRLAMP sensing cartridge based on direct sensing of the bulk refractive index.     

Detection of Staphylococcus aureus enterotoxin B at femtomolar levels with a miniature integrated two-channel surface plasmon resonance (SPR) sensor

Surface plasmon resonance (SPR) biosensors offer the capability for continuous real-time monitoring. The commercial instruments available have been large in size, expensive, and not amenable to field applications. We report here an SPR sensor system based on a prototype two-channel system similar to the single channel Spreeta devices. This system is an ideal candidate for field use. The two-channel design provides a reference channel to compensate for bulk refractive index (RI), non-specific binding and temperature variations. The SPR software includes a calibration function that normalizes the response from both channels, thus enabling accurate referencing. In addition, a temperature-controlled enclosure utilizing a thermo-electric module based on the Peltier effect provides the temperature stability necessary for accurate measurements of RI. The complete SPR sensor system can be powered by a 12V battery. Pre-functionalized, disposable, gold-coated thin glass slides provide easily renewable sensor elements for the system. Staphylococcus aureus enterotoxin B (SEB), a small protein toxin was directly detectable at sub-nanomolar levels and with amplification at femtomolar levels. A regeneration procedure for the sensor surface allowed for over 60 direct detection cycles in a 1-month period.     

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.     

Detection of gastric carcinoma-associated antigen MG7-Ag in human sera using surface plasmon resonance sensor

Background: MG7-Ag is a kind of gastric cancer-specific tumor-associated antigen and has been investigated to serve as a marker of gastric cancer for early diagnosis. Methods: Surface plasmon resonance (SPR) sensor was used for the detection of MG7-Ag in the sera of gastric cancer patients to develop an innovative, simple and rapid assay method for early diagnosis. The specific monoclonal MG7 antibodies were used as capture and detection receptors which were immobilized on the surface of SPR sensor chips for MG7-Ag identification in the human sera. The measurements include 9 cases of gastric cancer patients and 2 cases of healthy blood donors and a MKN45 cancer cell lysate solution sample for positive control. Results: The binding of MG7-Ag onto the sensor surface was observed from SPR spectra. The sera of most gastric cancer patients revealed much higher expression level of MG7-Ag than healthy human sera did in SPR measurement. Conclusion: The initial results demonstrate that the SPR biosensor has the potential for its application in the early diagnosis of gastric cancer. However, more tests need to be done to confirm the detection limitation and the criterion for cancer risk evaluation in early diagnosis.     

Evaluation of two- and three-dimensional streptavidin binding platforms for surface plasmon resonance spectroscopy studies of DNA hybridization and protein-DNA binding

Surface plasmon resonance (SPR) spectroscopy has been used to study DNA assembly, DNA hybridization, and protein-DNA interactions on two streptavidin (SA) sensor chips. On one chip, SA molecules are immobilized on a biotin-exposed surface, forming an ordered two-dimensional (2D) SA monolayer. The other chip, BIAcore's SA chip, contains SA molecules immobilized within a three-dimensional (3D) carboxylated dextran matrix. Compared to the 2D chip, the 3D SA matrix allows for a slower immobilization rate of biotinylated DNA due to diffusion limitation in the dextran matrix, but with twice the amount of the immobilized DNA due to the greater number of reactive sites, which in turn enables a higher sensitivity for DNA hybridization detection. Interestingly, having a greater DNA probe dispersion in the 3D matrix does not induce a higher DNA hybridization efficiency. In a study of protein binding to immobilized DNA (estrogen receptor to estrogen response elements), aiming at assessing the DNA sequence dependent protein binding behavior, the 2D and 3D chips produce different binding characteristics. On the 2D chip, the protein binding exhibits a better selectivity to the specific sequences, regardless of binding stringency (e.g. salt concentration), whereas on the 3D chip, the liquid handling system needs to be optimized in order to minimize transport limitations and to detect small affinity differences. Through this study we demonstrate that the physicochemical structure of SPR chips affects the apparent binding behaviors of biomolecules. When interpreting SPR binding curves and selecting a sensor chip, these effects should be taken into account.     

Immobilisation of DNA probes for the development of SPR-based sensing

An immobilisation procedure based on the direct coupling of thiol-derivatised oligonucleotide probes to bare gold sensor surfaces has been used for DNA sensing applications. The instrumentation used relies on surface plasmon resonance (SPR) transduction; in particular the commercially available instruments BIACORE X and SPREETA, have been employed in this study. The performances of the SPR-based DNA sensors resulting from direct coupling of thiol-derivatised DNA probes onto gold chips, have been studied in terms of the main analytical parameters, i.e. selectivity, sensitivity, reproducibility, analysis time, etc. A comparison between the thiol-derivatised immobilisation approach and a reference immobilisation method, based on the coupling of biotinylated oligonucleotide probes onto a streptavidin coated dextran sensor surface, using synthetic complementary oligonucleotides has been discussed. Finally, a denaturation method to obtain ssDNA ready for hybridisation analysis has been applied to polymerase chain reaction (PCR) amplified samples, for the detection of genetically modified organisms (GMOs).     

Integrated Terahertz Surface Plasmon Resonance on Polyvinylidene Fluoride Layer for the Profiling of Fluid Reflectance Spectra

We design terahertz (THz) surface-plasmon-resonance (SPR) sensors using a ferroelectric polyvinylidene fluoride (PVDF) thin layer for biological sensing. The reflectivity properties based on SPR are described using transfer matrix method (TMM) and numerically simulated using finite-difference time domain (FDTD) method. The sensing characteristics of the structure are systematically analyzed through the examination of the reflectivity spectrum. The results reveal that the pronounced SPR resonance peak has quasi-linear relationship with the refractive index variation of the material under investigation. Through analyzing and optimizing the structural parameters of the THz SPR sensor, we achieved the theoretical value of the refractive index detection sensitivity as high as 0.393 THz/unit change of refractive index (RIU) for a 20-μm-thick liquid sample with a 10-μm PVDF layer. This work shows great promise toward realizing a THz SPR sensor with high sensitivity for identifying the signatures of biological fluid sample.     

Collection mode surface plasmon fibre sensors: a new biosensing platform

Sensors based on surface plasmon resonance (SPR) allow rapid, label-free, highly sensitive detection, and indeed this phenomenon underpins the only label-free optical biosensing technology that is available commercially. In these sensors, the existence of surface plasmons is inferred indirectly from absorption features that correspond to the coupling of light into a thin metallic film. Although SPR is not intrinsically a radiative process, when the metallic coating which support the plasmonic wave exhibits a significant surface roughness, the surface plasmon can itself couple to the local photon states, and emit light. Here we show that using silver coated optical fibres, this novel SPR transducing mechanism offers significant advantages compare to traditional reflectance based measurements such as lower dependency on the metallic thickness and higher signal to noise ratio. Furthermore, we show that more complex sensor architectures with multiple sensing regions scattered along a single optical fibre enable multiplexed detection and dynamic self referencing of the sensing signal. Moreover, this alternative approach allows to combine two different sensing technologies, SPR and fluorescence sensing within the same device, which has never been demonstrated previously. As a preliminary proof of concept of potential application, this approach has been used to demonstrate the detection of the seasonal influenza A virus.     

Surface plasmon resonance in protein-membrane interactions

Surface plasmon resonance (SPR) has become one of the most important techniques for studying macromolecular interactions. The most obvious advantages of SPR over other techniques are: direct and rapid determination of association and dissociation rates of binding process, no need for labelling of protein or lipids, and small amounts of sample used in the assay (often nM concentrations of proteins). In biochemistry, SPR is used mainly to study protein-protein interactions. On the other hand, protein-membrane interactions, although crucial for many cell processes, are less well studied. Recent advances in the preparation of stable membrane-like surfaces and the commercialisation of sensor chips has enabled widespread use of SPR in protein-membrane interactions. One of the most popular is Biacore's L1 sensor chip that allows capture of intact liposomes or even subcellular preparations. Lipid specificity of protein-membrane interactions can, therefore, be easily studied by manipulating the lipid composition of the immobilised membrane. The number of published papers has increased steadily in the last few years and the examples include domains or proteins that participate in cell signalling, pore-forming proteins, membrane-interacting peptides, coagulation factors, enzymes, amyloidogenic proteins, prions, etc. This paper gives a brief overview of different membrane-mimetic surfaces that can be prepared on the surface of SPR chips, properties of liposomes on the surface of L1 chips and some selected examples of protein-membrane interactions studied with such system.     

Highly Sensitive Plasmonic Refractive Index Sensor Based on Dual D-Shaped Photonic Crystal Fiber with Aluminum Nitride-Silver Films

A dual-core and dual D-shaped photonic crystal fiber (PCF)-based surface plasmon resonance (SPR) sensor with silver and aluminum nitride (AlN) films is designed. The distribution characteristics of the electromagnetic fields of core and plasmon modes, as well as the sensing properties, are numerically studied by finite element method (FEM). The structure parameters of the designed sensor are optimized by the optical loss spectrum. The results show the resonance wavelength variation of 489 nm for the refractive index (RI) range of 1.36?~?1.42. In addition, a maximum wavelength sensitivity of 13,400 nm/RIU with the corresponding RI resolution of 7.46?×?10^?6 RIU is obtained in the RI range of 1.41?~?1.42. The proposed sensor with the merits of high sensitivity, low cost, and simple structure has a wide application in the fields of RI sensing, such as hazardous gas detection, environmental monitoring, and biochemical analysis.

     

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.