Marinus A. Otte

Trends and challenges of refractometric nanoplasmonic biosensors: a review.

Motivated by potential benefits such as sensor miniaturization, multiplexing opportunities and higher sensitivities, refractometric nanoplasmonic biosensing has profiled itself in a short time span as an interesting alternative to conventional Surface Plasmon Resonance (SPR) biosensors. This latter conventional sensing concept has been subjected during the last decades to strong commercialization, thereby strongly leaning on well-developed thin-film surface chemistry protocols. Not surprisingly, the examples found in literature based on this sensing concept are generally characterized by extensive analytical studies of relevant clinical and diagnostic problems. In contrast, the more novel Localized Surface Plasmon Resonance (LSPR) alternative finds itself in a much earlier, and especially, more fundamental stage of development. Driven by new fabrication methodologies to create nanostructured substrates, published work typically focuses on the novelty of the presented material, its optical properties and its use - generally limited to a proof-of-concept - as a label-free biosensing scheme. Given the different stages of development both SPR and LSPR sensors find themselves in, it becomes apparent that providing a comparative analysis of both concepts is not a trivial task. Nevertheless, in this review we make an effort to provide an overview that illustrates the progress booked in both fields during the last five years. First, we discuss the most relevant advances in SPR biosensing, including interesting analytical applications, together with different strategies that assure improvements in performance, throughput and/or integration. Subsequently, the remaining part of this work focuses on the use of nanoplasmonic sensors for real label-free biosensing applications. First, we discuss the motivation that serves as a driving force behind this research topic, together with a brief summary that comprises the main fabrication methodologies used in this field. Next, the sensing performance of LSPR sensors is examined by analyzing different parameters that can be invoked in order to quantitatively assess their overall sensing performance. Two aspects are highlighted that turn out to be especially important when trying to maximize their sensing performance, being (1) the targeted functionalization of the electromagnetic hotspots of the nanostructures, and (2) overcoming inherent negative influence that stem from the presence of a high refractive index substrate that supports the nanostructures. Next, although few in numbers, an overview is given of the most exhaustive and diagnostically relevant LSPR sensing assays that have been recently reported in literature, followed by examples that exploit inherent LSPR characteristics in order to create highly integrated and high-throughput optical biosensors. Finally, we discuss a series of considerations that, in our opinion, should be addressed in order to bring the realization of a stand-alone LSPR biosensor with competitive levels of sensitivity, robustness and integration (when compared to a conventional SPR sensor) much closer to reality.

Identification of the Optimal Spectral Region for Plasmonic and Nanoplasmonic Sensing

We present a theoretical and experimental study involving the sensing characteristics of wavelength-interrogated plasmonic sensors based on surface plasmon polaritons (SPP) in planar gold films and on localized surface plasmon resonances (LSPR) of single gold nanorods. The tunability of both sensing platforms allowed us to analyze their bulk and surface sensing characteristics as a function of the plasmon resonance position. We demonstrate that a general figure of merit (FOM), which is equivalent in wavelength and energy scales, can be employed to mutually compare both sensing schemes. Most interestingly, this FOM has revealed a spectral region for which the surface sensitivity performance of both sensor types is optimized, which we attribute to the intrinsic dielectric properties of plasmonic materials. Additionally, in good agreement with theoretical predictions, we experimentally demonstrate that, although the SPP sensor offers a much better bulk sensitivity, the LSPR sensor shows an approximately 15% better performance for surface sensitivity measurements when its FOM is optimized. However, optimization of the substrate refractive index and the accessibility of the relevant molecules to the nanoparticles can lead to a total 3-fold improvement of the FOM in LSPR sensors.

Direct detection of protein biomarkers in human fluids using site-specific antibody immobilization strategies.

Design of an optimal surface biofunctionalization still remains an important challenge for the application of biosensors in clinical practice and therapeutic follow-up. Optical biosensors offer real-time monitoring and highly sensitive label-free analysis, along with great potential to be transferred to portable devices. When applied in direct immunoassays, their analytical features depend strongly on the antibody immobilization strategy. A strategy for correct immobilization of antibodies based on the use of ProLinker™ has been evaluated and optimized in terms of sensitivity, selectivity, stability and reproducibility. Special effort has been focused on avoiding antibody manipulation, preventing nonspecific adsorption and obtaining a robust biosurface with regeneration capabilities. ProLinker™-based approach has demonstrated to fulfill those crucial requirements and, in combination with PEG-derivative compounds, has shown encouraging results for direct detection in biological fluids, such as pure urine or diluted serum. Furthermore, we have implemented the ProLinker™ strategy to a novel nanoplasmonic-based biosensor resulting in promising advantages for its application in clinical and biomedical diagnosis.

Highly sensitive dendrimer-based nanoplasmonic biosensor for drug allergy diagnosis.

A label-free biosensing strategy for amoxicillin (AX) allergy diagnosis based on the combination of novel dendrimer-based conjugates and a recently developed nanoplasmonic sensor technology is reported. Gold nanodisks were functionalized with a custom-designed thiol-ending-polyamido-based dendron (d-BAPAD) peripherally decorated with amoxicilloyl (AXO) groups (d-BAPAD-AXO) in order to detect specific IgE generated in patients serum against this antibiotic during an allergy outbreak. This innovative strategy, which follows a simple one-step immobilization procedure, shows exceptional results in terms of sensitivity and robustness, leading to a highly-reproducible and long-term stable surface which allows achieving extremely low limits of detection. Moreover, the viability of this biosensor approach to analyze human biological samples has been demonstrated by directly analyzing and quantifying specific anti-AX antibodies in patients serum without any sample pretreatment. An excellent limit of detection (LoD) of 0.6ng/mL (i.e. 0.25kU/L) has been achieved in the evaluation of clinical samples evidencing the potential of our nanoplasmonic biosensor as an advanced diagnostic tool to quickly identify allergic patients. The results have been compared and validated with a conventional clinical immunofluorescence assay (ImmunoCAP test), confirming an excellent correlation between both techniques. The combination of a novel compact nanoplasmonic platform and a dendrimer-based strategy provides a highly sensitive label free biosensor approach with over two times better detectability than conventional SPR. Both the biosensor device and the carrier structure hold great potential in clinical diagnosis for biomarker analysis in whole serum samples and other human biological samples.

Guiding light in monolayers of sparse and random plasmonic meta-atoms.

Encouraged by the capacity of surface plasmons to confine and propagate electromagnetic fields, waveguiding concepts have been developed, including combinations of continuous metal films or ordered arrays of metal nanoparticles. So far, waveguiding in the latter systems has been based on near-field or diffractive coupling. Herein, we show that monolayers of sparse and disordered gold nanoparticles support a novel transverse-electric guided mode that, contrary to previous work, relies on the strong enhancement of the polarizability upon excitation of the nanoparticle LSPR, creating an effective refractive index sufficiently high to support light guidance over a large range of frequencies. Excitation of this guided mode offers interesting nanophotonics features and applications such as a tunable total absorption spectral band, attractive for light harvesting applications, or the generation of a large amplification of the sensitivity to changes of refractive index accompanied with striking enhancement of the limit of detection in real biosensing experiments.

Surface plasmon resonance biosensors for highly sensitive detection in real samples

In this work we summarize the main results obtained with the portable surface plasmon resonance (SPR) device developed in our group (commercialised by SENSIA, SL, Spain), highlighting its applicability for the real-time detection of extremely low concentrations of toxic pesticides in environmental water samples. In addition, we show applications in clinical diagnosis as, on the one hand, the real-time and label-free detection of DNA hybridization and single point mutations at the gene BRCA-1, related to the predisposition in women to develop an inherited breast cancer and, on the other hand, the analysis of protein biomarkers in biological samples (urine, serum) for early detection of diseases. Despite the large number of applications already proven, the SPR technology has two main drawbacks: (i) not enough sensitivity for some specific applications (where pM-fM or single-molecule detection are needed) (ii) low multiplexing capabilities. In order solve such drawbacks, we work in several alternative configurations as the Magneto-optical Surface Plasmon Resonance sensor (MOSPR) based on a combination of magnetooptical and ferromagnetic materials, to improve the SPR sensitivity, or the Localized Surface Plasmon Resonance (LSPR) based on nanostructures (nanoparticles, nanoholes,...), for higher multiplexing capabilities.

Tailored Height Gradients in Vertical Nanowire Arrays via Mechanical and Electronic Modulation of Metal-Assisted Chemical Etching

In current top-down nanofabrication methodologies the design freedom is generally constrained to the two lateral dimensions, and is only limited by the resolution of the employed nanolithographic technique. However, nanostructure height, which relies on certain mask-dependent material deposition or etching techniques, is usually uniform, and on-chip variation of this parameter is difficult and generally limited to very simple patterns. Herein, a novel nanofabrication methodology is presented, which enables the generation of high aspect-ratio nanostructure arrays with height gradients in arbitrary directions by a single and fast etching process. Based on metal-assisted chemical etching using a catalytic gold layer perforated with nanoholes, it is demonstrated how nanostructure arrays with directional height gradients can be accurately tailored by: (i) the control of the mass transport through the nanohole array, (ii) the mechanical properties of the perforated metal layer, and (iii) the conductive coupling to the surrounding gold film to accelerate the local electrochemical etching process. The proposed technique, enabling 20-fold on-chip variation of nanostructure height in a spatial range of a few micrometers, offers a new tool for the creation of novel types of nano-assemblies and metamaterials with interesting technological applications in fields such as nanophotonics, nanophononics, microfluidics or biomechanics.

Spatial Distribution of Optical Near-Fields in Plasmonic Gold Sphere Segment Voids

We present a comprehensive experimental and computational study on the electromagnetic field distribution in sphere segment void arrays. Surface plasmon polaritons can be excited in these void arrays, resulting in greatly enhanced electromagnetic fields. With the scanning near-field optical microscope (SNOM) we are able to measure the electromagnetic field distribution at the sample surface. For this purpose, an array of relatively large voids with a sphere diameter of 900 nm was fabricated, allowing for an easy access of the scanning glass-fibre tip and yielding very detailed scans. Complementary, finite-difference time-domain (FDTD) calculations on a complete void array have been performed and compared with the SNOM intensity maps and experimental reflectivity data. We show in a direct way both the existence of extended and localised modes in the Au void array for three different void depths. We also show and discuss the changes that the modes undergo for the different void depths and excitation wavelengths. Moreover, since the simulations were performed for two different void geometries, one containing perfectly spherical void surfaces and another more realistic one, which considers the presence of interstitial wall holes and other imperfections, as observed in scanning electron micrographs, we were able to determine by comparison with the experiment under which conditions an array of idealised sphere segment voids is a meaningful model. This demonstrates that both SNOM and FDTD simulations are powerful tools for understanding the plasmonic response of metallic nanostructures, thus enabling, for instance, a design for applications in ultra-sensitive optical detection.