ssDNA aptamer-based surface plasmon resonance biosensor for the detection of retinol binding protein 4 for the early diagnosis of type 2 diabetes

Retinol binding protein 4 (RBP4) is a useful biomarker in the diagnosis of type 2 diabetes since its level in the serum is higher in insulin-resistant states. Accurate measurement of the serum RBP4 levels is hampered by conventional immunologic methods, such as enzyme-linked immunosorbent assay (ELISA). In this study, therefore, we have developed an aptamer-based surface plasmon resonance (SPR) biosensor that can be used to sense for RBP4 in serum samples. A single-stranded DNA (ssDNA) aptamer that showed high affinity (Kd = 0.2 +/- 0.03 microM) and specificity to RBP4 was selected. This RBP4-specific aptamer was immobilized on a gold chip and used in a label-free RBP4 detection using SPR. Analysis of RBP4 in artificial serum using SPR was compared with ELISA and Western blot analysis. Our results indicated that the RBP4-specific aptamer-based SPR biosensor gave better dose-dependent responses and was more sensitive than ELISA assays. As such, this RBP4 aptamer-based SPR biosensor can be potentially used to monitor the RBP4 levels within the serum as an indicator of type 2 diabetes.     

Mass sensitivity calculation of the protein layer using love wave SAW biosensor

Love waves, a variety of surface acoustic waves (SAWs), can be used to detect very small biological surface interactions and so have a wide range of potential applications. To demonstrate the practicality of a Love wave SAW biosensor, we fabricated a 155-MHz Love wave SAW biosensor and compared it with a commercial surface Plasmon resonance (SPR) using glycerol-water solution with known densities and viscosities to calibrate the response signals of the biosensors. And the mass per unit area of anti-mouse IgG bound with protein G onto the sensitive layer of the biosensor was calculated on the basis of the calibration result. The sensitivity of the Love wave SAW biosensor was the same as or greater than that of the SPR biosensor. Furthermore, the Love wave SAW biosensor was capable of measuring a much wider range of viscosities than the SPR biosensor. Although the operating principle of the Love wave SAW biosensor is completely different from that of the SPR biosensor, the subtle changes in the viscoelastic properties of the biological layer that accompany biological binding reactions on the sensitive layer can be monitored and measured in the same ways as with the SPR biosensor.     

Label-free detection of proteins in crude cell lysate with antibody arrays by a surface plasmon resonance imaging technique

We established a label-free method of measuring proteins in crude cell lysate using antibody arrays and surface plasmon resonance (SPR) imaging. The refractivity of the running buffer was adjusted with that of the lysate to overcome the bulk effect. The chemistries of the fabricated arrays were investigated to reduce nonspecific adsorption on the array surface. We found that the hydrophilicity of the poly(ethylene glycol) moiety and lower electrostatic charge on the surface provided a specific measurement of antigen-antibody interaction. We validated the system by measuring the expression of eight proteins in the mouse brain and comparing the results to those by conventional Western blotting. The detection limit of the antibody array was approximately 30 ng/mL in crude cell lysate, on the same order as that of previous SPR research. The system enabled quick, label-free, and high-throughput analysis of abundant proteins with minimal sample volume ( approximately 200 muL). It is expected that our SPR antibody array will be applicable for direct protein expression profiling of cell lysate, as well as for cell phenotyping, food analysis, discovery of new biomarkers, and immunological disease diagnostics.     

Improved sensitivity of wavelength-modulated surface plasmon resonance biosensor using gold nanorods

In this report, gold nanorods (GNRs) were used to enhance the sensitivity of the wavelength-modulated surface plasmon resonance (SPR) biosensor. The GNRs were designed and fabricated through seed-medicated growth and surface activation by a layer of a weak polyelectrolyte, poly(acrylic acid) for the attaching antibody. Rabbit anti-goat IgG was immobilized on GNRs, and sandwich assays were carried out to detect goat IgG using a wavelength-modulated SPR biosensor. The detection sensitivity of the nanorod-conjugated antibody is 25-100 times more sensitive than the SPR biosensor without GNRs. Drastic sensitivity enhancement, owing to the electromagnetic interaction between the nanotag and the sensing film, was maximized using the longitudinal plasmonic resonance of the GNRs. GNRs could significantly enhance the sensitivity of the SPR biosensor, and the maximum enhancement effect can be achieved when the longitudinal SPR peak wavelength of GNRs functionally matches the surface plasmon wavelength.     

Label-free detection of leptin antibody-antigen interaction by using LSPR-based optical biosensor

In the recent research, the development of optical biosensing devices has been focused on finding new method and technologies to exploit the optical properties of noble metal nanostructure, especially localized surface plasmon resonance (LSPR). In this study, we fabricated a LSPR-based label-free optical biosensor with the multi-spot gold-capped nanoparticle array (MG-NPA) biochip based on the deposition of a thin gold (Au) film on the silica nanoparticles layer with the simple process. The MG-NPA biochip used the silica nanoparticles as the core and a thin Au film as a shell on the surface. This structure can excite the LSPR signal easily with the high reproducibility. The anti-leptin antibody was immobilized on the surface of MG-NPA biochip, which could recognize only leptin antigen. The leptin antibody-antigen interaction was performed by the introduction of different concentration (1 pg/mL-100 microg/mL) of leptin antigen solutions for 1 h. The detection limit was found to be 100 pg/mL by using the anti-leptin antibody immobilized MG-NPA biochip. This LSPR-based label-free optical biosensor employing the MG-NPA biochip brings several advantages such as low cost, easy to fabricate, using a simple optical system and can be applied in a wide immunoassay with the similar antibody-antigen model.     

In situ biosensing with a surface plasmon resonance fiber grating aptasensor

Surface plasmon resonance (SPR) biosensors prepared using optical fibers can be used as a cost-effective and relatively simple-to-implement alternative to well established biosensor platforms for monitoring biomolecular interactions in situ or possibly in vivo. The fiber biosensor presented in this study utilizes an in-fiber tilted Bragg grating to excite the SPR on the surface of the sensor over a large range of external medium refractive indices, with minimal cross-sensitivity to temperature and without compromising the structural integrity of the fiber. The label-free biorecognition scheme used demonstrates that the sensor relies on the functionalization of the gold-coated fiber with aptamers, synthetic DNA sequences that bind with high specificity to a given target. In addition to monitoring the functionalization of the fiber by the aptamers in real-time, the results also show how the fiber biosensor can detect the presence of the aptamer's target, in various concentrations of thrombin in buffer and serum solutions. The findings also show how the SPR biosensor can be used to evaluate the dissociation constant (K(d)), as the binding constant agrees with values already reported in the literature.     

A colorimetric gold nanoparticle sensor to interrogate biomolecular interactions in real time on a surface

This paper presents a new label-free optical method to study biomolecular interactions in real time at the surface of an optically transparent substrate. The method relies on the change in the absorbance spectrum of a self-assembled monolayer of colloidal gold on glass, as a function of biomolecular binding to the surface of the immobilized colloids. Using this approach, we demonstrate proof of principle of a label-free optical biosensor to quantify biomolecular interactions in real time on a surface in a commercially available UV-visible spectrophotometer and of a colorimetric end-point assay using an optical scanner. The spectrophotometric sensor shows concentration-dependent binding and a detection limit of 16 nM for streptavidin. The sensor is easy to fabricate, is reproducible in its performance, has minimal technological requirements, namely, the availability of an UV-visible spectrophotometer or an optical scanner, and will enable high-throughput screening of biomolecular interactions in real time in an array-based format.     

Development of surface plasmon resonance mass spectrometry array platform

Protein microarrays are the format of choice for high-throughput, high-content protein interaction analysis. In most of the array formats, reporter molecules are used in multistep detection of the protein interactions. Among the few existing label-free detection approaches, surface plasmon resonance (SPR) and mass spectrometry (MS) stand out as most promising for utilization in protein microarrays, albeit both have been used only sporadically for high-content protein arrays. Shown here for the first time is the combination of SPR and MS detection on a single high-content protein microarray. Antibodies to five human plasma proteins were arrayed in a 10 x 10 spot arrangement on a chemically activated gold-coated glass chip. Binding of proteins to their corresponding antibodies was monitored via SPR imaging across the entire surface of the chip. Following protein affinity retrieval, the chip was overlaid with MALDI matrix and MS analyzed, producing protein-specific mass spectra from distinct spots on the array. The SPR-MS dual detection is well suited for high-content protein microarrays and comprehensive protein analysis-from quantitative assessment of the protein concentration to detection of structural protein variants arising from genetic variations and postexpression processing.     

Single-layer graphene-based surface plasmon resonance sensor with dynamic evanescent field enhancement for biomarker study

ABSTRACT

In this work, we have developed a kind of single-layer graphene-based surface plasmon resonance (SLG-SPR) biosensor to detect C-reactive protein (CRP) and Prostate-specific antigen (PSA). In the experiment of testing CPR, the results obtained revealed that the changes in resonance wavelength of SLG-SPR biosensors are higher than that of the gold-film based SPR (Au-SPR) biosensors. Moreover, for the experiment of testing PSA, due to the dynamic evanescent field enhancement produced by a strong electric field coupling between the localized SPR (LSPR) of AuNPs and SPR of single-layer graphene-based film (SLG-film) that further amplify the evanescent field signal. We verified the SLG-SPR biosensors exhibited higher sensitivity than the Au-SPR biosensors and the SLG-SPR biosensor exceeded the traditional biosensor detection limit. Accordingly, the SLG-SPR biosensor based on dynamic optical enhancement can realize high sensitivity detection of low concentration biomarkers and can be applied to most of the trace biomarkers in theory.     

Electrochemical DNA biosensor based on conducting polyaniline nanotube array

Most of the recent developments in ultrasensitive detection of nucleic acid are based on the gold nanoparticles and carbon nanotubes as a medium of signal amplification. Here, we present an ultrasensitive electrochemical nucleic acid biosensor using the conducting polyaniline (PANI) nanotube array as the signal enhancement element. The PANI nanotube array of a highly organized structure was fabricated under a well-controlled nanoscale dimension on the graphite electrode using a thin nanoporous layer as a template, and 21-mer oligonucleotide probes were immobilized on these PANI nanotubes. In comparison with gold nanoparticle- or carbon nanotube-based DNA biosensors, our PANI nanotube array-based DNA biosensor could achieve similar sensitivity without catalytic enhancement, purification, or end-opening processing. The electrochemical results showed that the conducting PANI nanotube array had a signal enhancement capability, allowing the DNA biosensor to readily detect the target oligonucleotide at a concentration as low as 1.0 fM (approximately 300 zmol of target molecules). In addition, this biosensor demonstrated good capability of differentiating the perfect matched target oligonucleotide from one-nucleotide mismatched oligonucleotides even at a concentration of 37.59 fM. This detection specificity indicates that this biosensor could be applied to single-nucleotide polymorphism analysis and single-mutation detection.     

Magnetic nanoparticle-enhanced biosensor based on grating-coupled surface plasmon resonance

A highly sensitive surface plasmon resonance (SPR) biosensor employing magnetic nanoparticle (MNP) assays is presented. In the reported approach, MNPs simultaneously served as "vehicles" for rapid delivery of target analyte from a sample to the sensor surface and as labels increasing the measured refractive index changes that are associated with the binding of target analyte. An optical setup based on grating-coupled surface plasmon resonance (GC-SPR) was used with a magnetic field gradient applied through the sensor chip for manipulating with MNPs on its surface. Iron oxide MNPs and a sensor surface with metallic diffraction grating were modified with antibodies that specifically recognize different epitopes of the analyte of interest. The sensitivity of the biosensor was investigated as a function of mass transport of the analyte to the sensor surface driven by diffusion (free analyte) or by the magnetic field gradient (analyte bound to MNPs). Immunoassay-based detection of β human chorionic gonadotropin (βhCG) was implemented to evaluate the sensitivity of the MNP-enhanced GC-SPR biosensor scheme. The results reveal that the sensitivity of βhCG detection was improved by 4 orders of magnitude compared with the regular SPR sensor with direct detection format, and a limit of detection below pM was achieved.     

Metallic film optimization in a surface plasmon resonance biosensor by the extended Rouard method

The extended Rouard method is applied to the computation of a multi-absorbing-layer system for the optimization of surface plasmon resonance (SPR) sensors. Specifically, the effect of the properties of a metallic layer on the shape of the reflectivity and sensitivity curve is demonstrated in the case of a Kretschmann configuration. This theoretical investigation allows us to establish the best optical properties of the metal to obtain a localized SPR, given the illuminating beam properties. Toward the development of a sensitive biosensor based on SPR, we quantify the changes in reflectivity of such an optical biosensor induced by the deposition of a nanometric biochemical film as a function of the metal film characteristics and the illumination operating conditions. The sensitivity of the system emphasizes the potential of such biophotonic technology using metallic multilayer configurations, especially with envisioned metamaterials.     

Sensitivity analysis of a nanowire-based surface plasmon resonance biosensor in the presence of surface roughness

We have investigated the effect of surface roughness on the sensitivity of conventional and nanowire-based surface plasmon resonance (SPR) biosensors. The theoretical research was conducted using rigorous coupled-wave analysis with Gaussian surface profiles of gold films determined by atomic force microscopy. The results suggest that, when surface roughness ranges near 1 nm, the sensitivity of a conventional SPR system is not significantly affected regardless of the correlation length. For a nanowire-based SPR biosensor, however, we have found that the sensitivity degrades substantially with decreasing correlation length. In particular, at a correlation length smaller than 100 nm, a random rough surface may induce destructive coupling between excited localized surface plasmons, which can lead to prominent reduction of sensitivity enhancement.     

Two-dimensional biosensor arrays based on surface plasmon resonance phase imaging

We present a biosensor design based on capturing the two-dimensional (2D) phase image of surface plasmon resonance (SPR). This 2D SPR imaging technique may enable parallel label-free detection of multiple analytes and is compatible with the microarray chip platform. This system uses our previously reported differential phase measurement approach, in which 2D phase maps obtained from the signal (P) and reference (S) polarizations are compared pixel by pixel. This technique greatly improves detection resolution as the subtraction step can eliminate measurement fluctuations caused by external disturbances as they essentially appear in both channels. Unlike conventional angular SPR systems, in which illumination from a range of angles must be used, phase measurement requires illumination from only one angle, thus making it well suited for 2D measurement. Also, phase-stepping introduced from a moving mirror provides the necessary modulation for accurate detection of the phase. In light of the rapidly increasing need for fast real-time detection, quantification, and identification of a range of proteins for various biomedical applications, our 2D SPR phase imaging technique should hold a promising future in the medical device market.     

Surface plasmon resonance imaging of biomolecular interactions on a grating-based sensor array

A surface plasmon resonance sensor array based upon a grating substrate was developed for the detection of biomolecular interactions. The substrate consisted of a gold grating prepared by wet chemical treatment of a commercial recordable compact disk. A custom-built floating pin microspotter was constructed to deliver solutions containing omega-functionalized linear alkanethiols to the grating surface and produce an array of sensor elements with different exposed functional end groups. This array platform can be used to study biomolecular interactions in a label-free, sensitive, and high-throughput format. To illustrate the performance of this device, a test protein (bovine serum albumin) was exposed to sensor elements containing an array of functionalized alkanethiols possessing either activated carboxylic acid-, amine-, or hydroxyl-terminated regions. Local changes in plasmon resonance were monitored in a fixed-angle imaging configuration. Plasmon images clearly distinguish the degree of protein attachment at the various surfaces. The molecular binding events on the grating were also confirmed by ellipsometry. This grating-based SPR imaging platform represents a simple and robust method for performing label-free, high-sensitivity, and high-throughput detection of biomolecular interactions.     

Characterization of Nanoporous Silicon-Based DNA Biosensor for the Detection of Salmonella Enteritidis

A biosensor for the detection of food-borne pathogens (Salmonella Enteritidis) was fabricated based on nanoporous silicon (NPS). P-type silicon wafers (100, 0.01 ) were anodized electrochemically in an electrochemical Teflon cell, containing ethanoic hydrofluoric acid solution to produce the porous layer on the silicon surface. The porous silicon surface was functionalized with DNA probes specific to the insertion element (Iel) gene of Salmonella Enteritidis. A biotin-streptavidin system was utilized to characterize the availability of the nanopores and the specificity of the DNA probe. Based on the electrical property of DNA, redox indicators and cyclic voltammetry were used for the characterization of the biosensor. Results showed that the DNA probe was specific to the target DNA, and the porous silicon-based biosensor had more active surface area and higher sensitivity (1 ng/mL) than the planar silicon-based biosensor. This simple, label-free porous silicon-based biosensor has potential applications in high-throughput detection of pathogens.     

Label-free detection of peptide nucleic acid-DNA hybridization using localized surface plasmon resonance based optical biosensor

The development of label-free optical biosensors for DNA and other biomolecules has the potential to impact life sciences as well as screening in medical and environmental applications. In this report, we developed a localized surface plasmon resonance (LSPR) based label-free optical biosensor based on a gold-capped nanoparticle layer substrate immobilized with peptide nucleic acids (PNAs). PNA probe was designed to recognize the target DNA related to tumor necrosis factor. The nanoparticle layer was formed on a gold-deposited glass substrate by the surface modified silica nanoparticles using silane-coupling reagent. The optical properties of gold-capped nanoparticle layer substrate were characterized through monitoring the changes in the absorbance strength, as the thickness of the biomolecular layer increased with hybridization. The detection of PNA-DNA hybridization with target oligonucleotides and PCR-amplified real samples were performed with a limit of detection value of 0.677 pM target DNA. Selective discrimination against a single-base mismatch was also achieved. Our LSPR-based biosensor with the gold-capped nanoparticle layer substrate is applicable to the design of biosensors for monitoring of the interaction of other biomolecules, such as proteins, whole cells, or receptors with a massively parallel detection capability in a highly miniaturized package.     

Nanoscale affinity chip interface for coupling inhibition SPR immunosensor screening with Nano-LC TOF MS

The on-line nanoscale coupling of a surface plasmon resonance (SPR)-based inhibition biosensor immunoassay (iBIA) for the screening of low molecular weight molecules with nano-liquid-chromatography electrospray ionization time-of-flight mass spectrometry (nano-LC ESI TOF MS) for identification is described. The interface is based on a reusable recovery chip (RC) that contains a nanoscale biosorbent composed of a hydrogel layer modified with antibodies raised against the analyte featuring the unique possibility of performance characterization using the SPR biosensor. Various hydrogel chemistries were evaluated, and the standard Biacore CM5 chip showed the highest capture capacity in combination with affinity-purified polyclonal antibodies. The procedure has four stages: the samples are prepared (1) and screened using a screening chip (SC) in the iBIA (2). Suspected noncompliant samples as being noncompliant are reinjected over the RC, and the analyte is captured at subnanogram level (3). The captured analyte is released, and the eluate is analyzed with nano-LC ESI TOF MS via a loop-type interface (4). The coupling of the technologies proved effective for screening enrofloxacin, a model compound, in incurred chicken muscle samples followed by identity confirmation in suspected noncompliant samples. Ciprofloxacin, a known metabolite of enrofloxacin, was identified as well in incurred chicken samples. This demonstrates the potential of the technologies coupled by means of a RC for the rapid screening and identification of known as well as unknown compounds. Finally, we demonstrate the feasibility of combining the two biosensor chips (SC and RC) with a robust chip-based nano-LC chip TOF MS system, thus providing a robust alternative triple-chip system.     

Surface plasmon resonance investigations of human epidermal growth factor receptor 2

This investigation utilizes surface plasmon resonance (SPR) spectroscopy to detect and quantify human epidermal growth factor receptor 2 (HER-2), an oncogene product that is over-expressed in some aggressive forms of breast cancer. Specifically, the HER-2 trans-membrane protein p185 and its extra cellular fragment p105 are analytes targeted in this work by using a gold-based biosensor slide on which an anti-HER-2 antibody has been immobilized by attachment to Protein G that is fixed to the gold film. A detection limit of > or =11 ng/mL for p185 resulted when trastuzumab was used as the anti-HER-2 antibody on the biosensor slide. Experiments with semi-purified p105 revealed that it binds weakly and reversibly to trastuzumab, therefore complicating its detection and quantification. Results of studies that reacted a 13-amino-acid peptide (PP13) from the HER-2 kinase domain with its specific antibody were critically different than p185 and p105 studies. Spectral analysis of the reflectivity at constant bulk buffer refractive index revealed a progressive negative SPR shift over time. A negative shift suggests that a loss of protein mass from the anti-PP13 antibody-Protein G biosensor is occurring. Several possibilities that may explain these negative SPR shifts are discussed.