A Low-Temperature Annealing Method for Alloy Nanostructures and Metasurfaces: Unlocking a Novel Degree of Freedom.

The material and exact shape of a nanostructure determine its optical response, which is especially strong for plasmonic metals. Unfortunately, only a few plasmonic metals are available, which limits the spectral range where these strong optical effects can be utilized. Alloying different plasmonic metals can overcome this limitation, at the expense of using a high-temperature alloying process, which adversely destroys the nanostructure shape. Here, a low-temperature alloying process is developed where the sample is heated at only 300 °C for 8 h followed by 30 min at 450 °C and Au-Ag nanostructures with a broad diversity of shapes, aspect ratios, and stoichiometries are fabricated. Energy-dispersive X-ray spectroscopy and X-ray photoelectron spectroscopy analyses confirm the homogeneous alloying through the entire sample. Varying the alloy stoichiometry tunes the optical response and controls spectral features, such as Fano resonances. Binary metasurfaces that combine nanostructures with different stoichiometries are fabricated using multiple-step electron-beam lithography, and their optical function as a hologram or a Fresnel zone plate is demonstrated at the visible wavelength of λ = 532 nm. This low-temperature annealing technique provides a versatile and cost-effective way of fabricating complex Au-Ag nanostructures with arbitrary stoichiometry.

Hybrid Metal-Dielectric Metasurfaces for Refractive Index Sensing.

Hybrid metal-dielectric nanostructures have recently gained prominence because they combine strong field enhancement of plasmonic metals and the several low-loss radiation channels of dielectric resonators, which are qualities pertaining to the best of both worlds. In this work, an array of such hybrid nanoantennas is successfully fabricated over a large area and utilized for bulk refractive index sensing with a sensitivity of 208 nm/RIU. Each nanoantenna combines a Si cylinder with an Al disk, separated by a SiO2 spacer. Its optical response is analyzed in detail using the multipoles supported by its subparts and their mutual coupling. The nanoantenna is further modified experimentally with an undercut in the SiO2 region to increase the interaction of the electric field with the background medium, which augments the sensitivity to 245 nm/RIU. A detailed multipole analysis of the hybrid nanoantenna supports our experimental findings.

High-quality single crystal diamond diffraction gratings fabricated by crystallographic etching.

We demonstrate a novel method for fabricating single crystal diamond diffraction gratings based on crystallographic etching that yields high-quality diffraction gratings from commercially available <100> diamond plates. Both V-groove and rectangular gratings were fabricated and characterised using scanning electron microscopy and atomic force microscopy, revealing angles of 57° and 87° depending on the crystal orientation, with mean roughness below Ra = 5 nm on the sidewalls. The gratings were also optically characterised, showing good agreement with simulated results. The fabrication method demonstrated in this contribution shows the way for manufacturing high-quality diamond diffractive components that surpass existing devices both in quality and manufacturability.

Photocatalytic ammonia production enhanced by a plasmonic near-field and hot electrons originating from aluminium nanostructures.

Ammonia production at room temperature and atmospheric pressure is in high demand to assist in energy saving and the protection of the environment worldwide, as well as to help reduce CO2 emissions. Recently, plasmonic nanomaterials have been frequently used for solar to chemical energy conversion, which has the potential to replace existing energy-intensive industrial processes. In our approach, plasmonic aluminium nanotriangles (AlNTs) were used to investigate the impact of plasmonic effects on photocatalytic ammonia production. Plasmonic near-field coupling to a semiconductor and hot electron generation from AlNTs were studied in detail through the use of electrochemical photocurrent measurements. A narrowband LED beam with a central wavelength at 365 nm was used to illuminate the AlNTs and their hot electron generation efficiency was estimated to be 2 × 10-4%, resulting in an ammonia production rate of 4 × 10-5 µM h-1 mW-1 cm-2, which corresponds to a quantum efficiency of 2.5 × 10-5%. In the case of plasmonic near-field coupling, AlNTs-embedded TiO2 demonstrates a charge-carrier generation efficiency of 2.7%, which is ∼2.3 times higher than that of bare TiO2. The ammonia production rate of AlNTs-TiO2 is 0.1 µM h-1 mW-1 cm-2 with a quantum efficiency of ∼0.06%, which corresponds to ∼2.4 times that of the rate demonstrated by bare TiO2 (0.04 µM h-1 mW-1 cm-2, quantum efficiency ∼ 0.025%). The obtained results confirm successful ammonia production through nitrogen splitting at room temperature and under atmospheric pressure. Moreover, according to the presented results, the use of plasmonic aluminium structures remarkably improves the ammonia production rate.

Electrochemical Sensor for Bilirubin Detection Using Screen Printed Electrodes Functionalized with Carbon Nanotubes and Graphene.

Practice oriented point-of-care diagnostics require easy-to-handle, miniaturized, and low-cost analytical tools. In a novel approach, screen printed carbon electrodes (SPEs), which were functionalized with nanomaterials, are employed for selective measurements of bilirubin, which is an important biomarker for jaundice. Multi-walled carbon nanotubes (MWCNT) and graphene separately deposited on SPEs provide the core of an electrochemical sensor for bilirubin. The electrocatalytic activity towards bilirubin oxidation (bilirubin to biliverdin) was observed at +0.25 V. In addition, a further peak corresponding to the electrochemical conversion of biliverdin into purpurin appeared at +0.48 V. When compared to MWCNT, the graphene type shows a 3-fold lower detection limit (0.3 ± 0.022 nM and 0.1 ± 0.018 nM, respectively), moreover, the graphene type exhibits a larger linear range (0.1-600 µM) than MWCNT (0.5-500 µM) with a two-fold better sensitivity, i.e., 30 nA µM-1 cm-2, and 15 nA µM-1 cm-2, respectively. The viability is validated through measurements of bilirubin in blood serum samples and the selectivity is ensured by inhibiting common interfering biological substrates using an ionic nafion membrane. The presented approach enables the design and implementation of low cost and miniaturized electrochemical sensors.

Label-Free Electrochemical Immunoassay for C-Reactive Protein.

C-reactive protein (CRP) is one of the most expressed proteins in blood during acute phase inflammation, and its minute level increase has also been recognized for the clinical diagnosis of cardio vascular diseases. Unfortunately, the available commercial immunoassays are labour intensive, require large sample volumes, and have practical limitations, such as low stability and high production costs. Hence, we have developed a simple, cost effective, and label-free electrochemical immunoassay for the measurement of CRP in a drop of serum sample using an immunosensor strip made up of a screen printed carbon electrode (SPE) modified with anti-CRP functionalized gold nanoparticles (AuNPs). The measurement relies on the decrease of the oxidation current of the redox indicator Fe3+/Fe2+, resulting from the immunoreaction between CRP and anti-CRP. Under optimal conditions, the present immunoassay measures CRP in a linear range from 0.4-200 nM (0.047-23.6 µg mL-1), with a detection limit of 0.15 nM (17 ng mL-1, S/N = 3) and sensitivity of 90.7 nA nM-1, in addition to a good reproducibility and storage stability. The analytical applicability of the presented immunoassay is verified by CRP measurements in human blood serum samples. This work provides the basis for a low-priced, safe, and easy-to-use point-of-care immunosensor assay to measure CRP at clinically relevant concentrations.

Strong Improvement of Long-Term Chemical and Thermal Stability of Plasmonic Silver Nanoantennas and Films.

Silver (Ag) nanostructures and thin films are advantageous plasmonic materials as they have significantly lower losses than gold (Au). Unfortunately, Ag nanostructures suffer from poor chemical and thermal stability, which limit their applications. Here, the mechanisms leading to the deterioration of Ag nanostructures are clarified. It is first shown that oxygen alone cannot oxidize Ag nanostructures. Then, experiments using X-ray photoelectron spectroscopy reveal that the amount of sulfur in ambient air is too low for efficient tarnishing of the Ag surface. Finally, water is found to be the most critical factor for the degradation of Ag nanostructures and thin films. At high relative humidity, adsorbed water forms a thin film enabling the migration of Ag ions at the Ag/air interface, which deteriorates the Ag nanostructures. A dehydration treatment is developed which alters the morphology of the deposited silver, leading to an improved chemical and thermal stability of the Ag nanostructures and films, which then remain stable for more than 14 weeks under ambient laboratory conditions. In addition, dehydration also improves significantly the root-mean-square roughness for Ag thin films deposited on a glass substrate.

Non-invasive continuous monitoring of pro-oxidant effects of engineered nanoparticles on aquatic microorganisms.

Engineered nanomaterials (ENMs) are key drivers for the development of highly sophisticated new technologies. As all new attainments, the rapidly increasing used of ENMs raise concerns about their safety for the environment and humans. There is growing evidence showing that if engineered nanomaterials are released into the environment, there is a possibility that they could cause harm to aquatic microorganisms. Among the divers effects triggering their toxicity the ability of ENMs to generate reactive oxygen species (ROS) capable of oxidizing biomolecules is currently considered a central mechanism of toxicity. Therefore, development of sensitive tools for quantification of the ROS generation and oxidative stress are highly sought. After briefly introducing ENMs-induced ROS generation and oxidative stress in the aquatic microorganisms (AMOs), this overview paper focuses on a new optical biosensor allowing sensitive and dynamic measurements of H2O2 in real-time using multiscattering enhanced absorption spectroscopy. Its principle is based on sensitive absorption measurements of the heme protein cytochrome c whose absorption spectrum alters with the oxidation state of constituent ferrous FeII and ferric FeIII. For biological applications cytochrome c was embedded in porous random media resulting in an extended optical path length through multiple scattering of light, which lowers the limit of detection to a few nM of H2O2. The sensor was also integrated in a microfluidic system containing micro-valves and sieves enabling more complex experimental conditions. To demonstrate its performance, abiotic absorption measurements of low concentrations of dye molecules and 10 nm gold particles were carried out achieving limits of detection in the low nM range. Other biologically relevant reactive oxygen species can be measured at sub-µM concentrations, which was shown for glucose and lactate through enzymatic reactions producing H2O2. In ecotoxicological investigations H2O2 excreted by aquatic microorganisms exposed to various stressors were measured. Pro-oxidant effects of nano-TiO2 and nano-CuO towards green alga Chlamydomonas reinhardtii were explored in various exposure media and under different light illuminations. Dynamics of Cd2+ induced effects on photosynthetic activity, sensitisation and recovery of cells of C. reinhardtii was also studied.

Full Color Generation Using Silver Tandem Nanodisks.

Plasmonic effects associated with metallic nanostructures have been widely studied for color generation. It became apparent that highly saturated and bright colors are hard to obtain, and very small nanostructures need to be fabricated. To address this issue, in this study, we employ metal-insulator-metal sandwich nanodisks that support enhanced in-phase electric dipole modes, which are blue-shifted with respect to a single metal disk. The blue shift enables the generation of short wavelength colors with larger nanostructures. The radiation modes hybridize with the Wood's anomaly in periodic structures, creating narrow and high-resonance peaks in the reflection and deep valleys in the transmission spectra, thus producing vivid complementary colors in both cases. Full colors can be achieved by tuning the radius of the nanodisks and the periodicity of the arrays. Good agreement between simulations and experiments is demonstrated and analyzed in CIE1931, sRGB, and HSV color spaces. The presented method has potential for applications in imaging, data storage, ultrafine displays, and plasmon-based biosensors.

Highly sensitive SERS analysis of the cyclic Arg-Gly-Asp peptide ligands of cells using nanogap antennas.

The cyclic RGD (cRGD) peptide ligands of cells have become widely used for treating several cancers. We report a highly sensitive analysis of c(RGDfC) using surface enhanced Raman spectroscopy (SERS) using single dimer nanogap antennas in aqueous environment. Good agreement between characteristic peaks of the SERS and the Raman spectra of bulk c(RGDfC) with its peptide's constituents were observed. The exhibited blinking of the SERS spectra and synchronization of intensity fluctuations, suggest that the SERS spectra acquired from single dimer nanogap antennas was dominated by the spectrum of single to a few molecules. SERS spectra of c(RGDfC) could be used to detect at the nanoscale, the cells' transmembrane proteins binding to its ligand. SERS of cyclic RGD on nanogap antenna.

Maximal absorption regime in random media.

Efficient optical energy transfer is key to many technologies, ranging from biosensing to photovoltaics. Here, for the first time we show that by introducing a random medium with appropriate filling factor, absorption in a specific volume can be maximized. Using both numerical simulations and an analytical diffusion model, we identify design rules to maximize absorption in the system with different geometrical and scattering properties. By combining a random medium with an open photonic cavity, we numerically demonstrate a 23-fold enhancement of the absorbed energy. We also show how absorption as high as 99% can be reached in a device as thin as 500 µm for normal incidence illumination. Finally, our data indicate that introducing a non-absorbing random medium into a light trapping system for thin solar cells can enhance absorption of energy by a factor of 2.2. This absorption enhancement, caused by the random medium, is broadband and wide-angle and can help design efficient solar cells, light trapping devices, biosensors and random lasers.

New insights into ROS dynamics: a multi-layered microfluidic chip for ecotoxicological studies on aquatic microorganisms.

Reactive oxygen species (ROS) play an important role in the life of every cell, including cellular defense and signaling mechanisms. Continuous and quantitative ROS sensing can provide valuable information about the cell state, but it remains a challenge to measure. Here, we introduce a multi-layered microfluidic chip with an integrated optical sensor for the continuous sensitive detection of extracellular hydrogen peroxide (H2O2), one of the most stable ROS. This platform includes hydraulically controlled microvalves and microsieves, which enable the precise control of toxicants and complex exposure sequences. In particular, we use this platform to study the dynamics of toxicity-induced ROS generation in the green microalga Chlamydomonas reinhardtii during short-term exposures, recovery periods, and subsequent re-exposures. Two cadmium-based toxicants with distinct internalization mechanisms are used as stress inducers: CdSe/ZnS quantum dots (Qdots) and ionic cadmium (Cd(2+)). Our results show the quantitative dynamics of ROS generation by the model microalga, the recovery of cell homeostasis after stress events and the cumulative nature of two consecutive exposures. The dissolution of quantum dots and its possible influence on toxicity and H2O2 depletion is discussed. The obtained insights are relevant from ecotoxicological and physiological perspectives.

Multiscattering-enhanced optical biosensor: multiplexed, non-invasive and continuous measurements of cellular processes.

The continuous measurement of uptake or release of biomarkers provides invaluable information for understanding and monitoring the metabolism of cells. In this work, a multiscattering-enhanced optical biosensor for the multiplexed, non-invasive, and continuous detection of hydrogen peroxide (H2O2), lactate and glucose is presented. The sensing scheme is based on optical monitoring of the oxidation state of the metalloprotein cytochrome c (cyt c). The analyte of interest is enzymatically converted into H2O2 leading to an oxidation of the cyt c. Contact microspotting is used to prepare nanoliter-sized sensing spots containing either pure cyt c, a mixture of cyt c with glucose oxidase (GOx) to detect glucose, or a mixture of cyt c with lactate oxidase (LOx) to detect lactate. The sensing spots are embedded in a multiscattering porous medium that enhances the optical signal. We achieve limits of detection down to 240 nM and 110 nM for lactate and glucose, respectively. A microfluidic embodiment enables multiplexed and crosstalk-free experiments on living organisms. As an example, we study the uptake of exogenously supplied glucose by the green algae Chlamydomonas reinhardtii and simultaneously monitor the stress-related generation of H2O2. This multifunctional detection scheme provides a powerful tool to study biochemical processes at cellular level.

A miniaturized electrochemical assay for homocysteine using screen-printed electrodes with cytochrome c anchored gold nanoparticles.

Determination of homocysteine (HcySH) is highly beneficial in human physiology and pathophysiology for diagnosis and prognosis of cardiovascular diseases (CVD). Unfortunately, the practicability of the existing methodologies for the determination of HcySH is limited in terms of sample requirements, preparation time and instrumentation cost. To overcome these limitations, we have developed a new miniaturized electrochemical assay for HcySH in which cytochrome c (cyt c) immobilized on gold nanoparticle (GNP) modified screen printed carbon electrode (SPE) is employed as a biosensing element. The electrochemical characterization of the biosensor (cyt c-GNP-SPE) shows quasi-reversible redox peaks at the potentials 0 and -0.2 V, confirming the cyt c binding. The methodology of quantification is based on the electrochemical oxidation of HcySH by the Fe(3+)/Fe(2+) crevice of cyt c, observed at a potential of +0.56 V. Using the amperometric technique, the detection limit of HcySH is found to be 0.3 ± 0.025 µM in the linear range between 0.4 µM and 700 µM, with a sensitivity of 3.8 ± 0.12 nA µM(-1) cm(-2). The practical application of the present assay is validated through the measurement of HcySH in blood plasma samples and the selectivity is ensured by eliminating the impact of the common interfering biological substrates using a Nafion membrane. This biosensor shows striking analytical properties of good repeatability, reproducibility (2.85% SD) and high stability (83% of its initial current response after 4 weeks). This work paves the way for cheap, efficient and reliable point-of-care biosensors for screening one of the major causes of deaths both in the developed and developing countries.

Portable oxidative stress sensor: dynamic and non-invasive measurements of extracellular H2O2 released by algae.

Reactive oxygen species (ROS) generated by aerobic organisms are essential for physiological processes such as cell signaling, apoptosis, immune defense and oxidative stress mechanisms. Unbalanced oxidant/antioxidant budgets are involved in many diseases and, therefore, the sensitive measurement of ROS is of great interest. Here, we present a new device for the real-time monitoring of oxidative stress by measuring one of the most stable ROS, namely hydrogen peroxide (H2O2). This portable oxidative stress sensor contains the heme protein cytochrome c (cyt c) as sensing element whose spectral response enables the detection of H2O2 down to a detection limit of 40 nM. This low detection limit is achieved by introducing cyt c in a random medium, enabling multiscattering that enhances the optical trajectory through the cyt c spot. A contact microspotting technique is used to produce reproducible and reusable cyt c spots which are stable for several days. Experiments in static and microfluidic regimes, as well as numerical simulations demonstrate the suitability of the cyt c/H2O2 reaction system for the real-time sensing of the kinetics of biological processes without H2O2 depletion in the measurement chamber. As an example, we detect the release of H2O2 from the green alga Chlamydomonas reinhardtii exposed to either 180 nM functionalized CdSe/ZnS core shell quantum dots, or to 10 mg/l TiO2 nanoparticles. The continuous measurement of extracellular H2O2 by this optical sensor with high sensitivity is a promising new means for real-time cytotoxicity tests, the investigation of oxidative stress and other physiological cell processes.

Multiscattering-enhanced absorption spectroscopy.

An original scheme for sensitive absorption measurements, particularly well-suited for low analyte concentrations, is presented. The technique is based on multiscattering-enhanced absorption spectroscopy (MEAS) and benefits from the advantages of conventional absorption spectroscopy: simplicity, rapidity, and low costs. The technique relies on extending the optical path through the sensing volume by suspending dielectric beads in the solution containing the analytes of interest, resulting in multiple scattering of light, which increases the optical path length through the sample. This way, a higher sensitivity and lower limit of detection, compared to those of conventional absorption spectroscopy, can be achieved. The approach is versatile and can be used for a broad variety of analytes. Here, it is applied to the detection of phenol red, 10 nm gold nanoparticles, and envy green fluorescence dye; the limit of detection is decreased by a factor of 7.2 for phenol red and a factor of 3.3 for nanoparticles and dye. The versatility of this approach is illustrated by its application in increasing the sensitivity of colorimetric detection with gold nanoparticle probes and a commercially available hydrogen peroxide bioassay. The influence of different parameters describing the scattering medium is investigated in detail experimentally and numerically, with very good agreement between the two. Those parameters can be effectively used to tailor the enhancement for specific applications and analytes.

Absorbance enhancement in microplate wells for improved-sensitivity biosensors.

A generic optical biosensing strategy was developed that relies on the absorbance enhancement phenomenon occurring in a multiple scattering matrix. Experimentally, inserts made of glass fiber membrane were placed into microplate wells in order to significantly lengthen the trajectory of the incident light through the sample and therefore increase the corresponding absorbance. Enhancement factor was calculated by comparing the absorbance values measured for a given amount of dye with and without the absorbance-enhancing inserts in the wells. Moreover, the dilution of dye in solutions with different refractive indices (RI) clearly revealed that the enhancement factor increased with the ΔRI between the membrane and the surrounding medium, reaching a maximum value (EF>25) when the membranes were dried. On this basis, two H2O2-biosensing systems were developed based on the biofunctionalization of the glass fiber inserts either with cytochrome c or horseradish peroxidase (HRP) and the analytical performances were systematically compared with the corresponding bioassay in solution. The efficiency of the absorbance-enhancement approach was particularly clear in the case of the cytochrome c-based biosensor with a sensitivity gain of 40 folds and wider dynamic range. Therefore, the developed strategy represents a promising way to convert standard colorimetric bioassays into optical biosensors with improved sensitivity.

Sensing the dynamics of oxidative stress using enhanced absorption in protein-loaded random media.

Reactive oxygen species play a key role in cell signalling and oxidative stress mechanisms, therefore, sensing their production by living organisms is of fundamental interest. Here we describe a novel biosensing method for extracellular detection of endogenous hydrogen peroxide (H2O2). The method is based on the enhancement of the optical absorption spectrum of the hemoprotein cytochrome c when loaded into a highly scattering random medium. Such a configuration enables, in contrast to existing techniques, non-invasive and dynamic detection of the oxidation of cyt c in the presence of H2O2 with unprecedented sensitivity. Dynamic information on the modification of the cell oxidative status of Chlamydomonas reinhardtii, an aquatic green algae, was obtained under oxidative stress conditions induced by the presence of trace concentrations of Cd(II). Furthermore, the dynamics of H2O2 production was investigated under different lighting conditions confirming the impact of Cd(II) on the photosynthetic activity of those phytoplanktonic cells.

Coupling strength can control the polarization twist of a plasmonic antenna.

The far-field polarization of the optical response of a plasmonic antenna can be tuned by subtly engineering of its geometry. In this paper, we develop design rules for nano antennas which enable the generation of circular polarized light via the excitation of circular plasmonic modes in the structure. Two initially orthogonal plasmonic modes are coupled in such a way that a rotational current is excited in the structure. Modifying this coupling strength from a weak to a strong regime controls the helicity of the scattered field. Finally, we introduce an original sensing approach that relies on the rotation of the incident polarization and demonstrates a sensitivity of 0.23 deg·nm(-1) or 33 deg·RIU(-1), related to changes of mechanical dimensions and the refractive index, respectively.

Biosensor based on chemically-designed anchorable cytochrome c for the detection of H2O2 released by aquatic cells.

A novel third generation biosensor was developed based on one-shot adsorption of chemically-modified cytochrome c (cyt c) onto bare gold electrodes. In contrast to the classic approach which consists of attaching cyt c onto an active self-assembled monolayer (SAM) priory chemisorbed on gold, here short-chain thiol derivatives (mercaptopropionic acid, MPA) were chemically introduced on cyt c protein shell via its lysine residues enabling the very fast formation (< 5 min) of an electroactive biological self-assembled monolayer (SAM) exhibiting a quasi-reversible electrochemical behavior and a fast direct electron transfer (ET). The heterogeneous ET rate constant was estimated to be k(s)=1600 s⁻¹, confirming that short anchors facilitate the ET via an efficient orientation of the heme pocket. In comparison, no ET was observed in the case of native and long-anchor (mercaptoundecanoic acid, MUA) modified cyt c directly adsorbed on gold. However, in both cases the ET was efficiently restored upon in-bulk generation of gold nanoparticles which acted as electron shuttles. This observation emphasizes that the lack of electroactivity might be caused by either an inappropriate orientation of the protein (native cyt c) or a critical distance (MUA-cyt c). Finally, the sensitivity of the bare gold electrode directly modified with MPA-cyt c to hydrogen peroxide (H2O2) was evaluated by amperometry and the so-made amperometric biosensor was able to perform real-time and non-invasive detection of endogeneous H2O2 released by unicellular aquatic microorganisms, Chlamydomonas reinhardtii, upon cadmium exposure.