Evaluation of Mesoporous TiO2 Layers as Glucose Optical Sensors.

Porous materials are currently the basis of many optical sensors because of their ability to provide a higher interaction between the light and the analyte, directly within the optical structure. In this study, mesoporous TiO2 layers were fabricated using a bottom-up synthesis approach in order to develop optical sensing structures. In comparison with more typical top-down fabrication strategies where the bulk constitutive material is etched in order to obtain the required porous medium, the use of a bottom-up fabrication approach potentially allows increasing the interconnectivity of the pore network, hence improving the surface and depth homogeneity of the fabricated layer and reducing production costs by synthesizing the layers on a larger scale. The sensing performance of the fabricated mesoporous TiO2 layers was assessed by means of the measurement of several glucose dilutions in water, estimating a limit of detection even below 0.15 mg/mL (15 mg/dL). All of these advantages make this platform a very promising candidate for the development of low-cost and high-performance optical sensors.

Performance improvement of a silicon nitride ring resonator biosensor operated in the TM mode at 1310†nm.

Silicon-based ring resonators have been demonstrated to be a key element to build lab-on-chip devices due to their ability to perform as label-free photonic sensors. In this work, we demonstrate photonic biosensing using silicon nitride ring resonators operated in the TM mode around 1310 nm wavelengths. Our results show that operating the devices using the TM mode results in an increased sensitivity in comparison with the typically used TE mode, while working at 1310 nm wavelengths compared to 1550 nm contributes to an increased quality factor. As a result, a reduction in the intrinsic limit of detection is achieved, indicating the suitability of TM modes in the 1310 nm regime for biosensing using integrated photonics.

Design of slow-light-enhanced bimodal interferometers using dimensionality reduction techniques.

Interferometers usually require long paths for the ever-increasing requirements of high-performance operation, which hinders the miniaturization and integration of photonic circuits into very compact devices. Slow-light based interferometers provide interesting advantages in terms of both compactness and sensitivity, although their optimization is computationally costly and inefficient, due to the large number of parameters to be simultaneously designed. Here we propose the design of slow-light-enhanced bimodal interferometers by using principal component analysis to reduce the high-dimensional design space. A low-dimensional hyperplane containing all optimized designs is provided and investigated for changes in the silicon core and cladding refractive index. As a result, all-dielectric single-channel interferometers as modulators of only 33 µm2 footprint and sensors with 19.2 × 103 2πrad/RIU·cm sensitivity values are reported and validated by 2 different simulation methods. This work allows the design and optimization of slow light interferometers for different applications by considering several performance criteria, which can be extended to other photonic structures.

Slow light bimodal interferometry in one-dimensional photonic crystal waveguides.

Strongly influenced by the advances in the semiconductor industry, the miniaturization and integration of optical circuits into smaller devices has stimulated considerable research efforts in recent decades. Among other structures, integrated interferometers play a prominent role in the development of photonic devices for on-chip applications ranging from optical communication networks to point-of-care analysis instruments. However, it has been a long-standing challenge to design extremely short interferometer schemes, as long interaction lengths are typically required for a complete modulation transition. Several approaches, including novel materials or sophisticated configurations, have been proposed to overcome some of these size limitations but at the expense of increasing fabrication complexity and cost. Here, we demonstrate for the first time slow light bimodal interferometric behaviour in an integrated single-channel one-dimensional photonic crystal. The proposed structure supports two electromagnetic modes of the same polarization that exhibit a large group velocity difference. Specifically, an over 20-fold reduction in the higher-order-mode group velocity is experimentally shown on a straightforward all-dielectric bimodal structure, leading to a remarkable optical path reduction compared to other conventional interferometers. Moreover, we experimentally demonstrate the significant performance improvement provided by the proposed bimodal photonic crystal interferometer in the creation of an ultra-compact optical modulator and a highly sensitive photonic sensor.

Continuous Detection of Increasing Concentrations of Thrombin Employing a Label-Free Photonic Crystal Aptasensor.

Thrombin generation is a complex and finely regulated pathway that provokes dynamical changes of thrombin concentration in blood when a vascular injury occurs. In order to characterize the initiation phase of such process, when thrombin concentration is in the nmol/L range, a label-free optical aptasensor is proposed here. This aptasensor combines a 1D photonic crystal structure consisting of a silicon corrugated waveguide with thrombin binding aptamers on its surface as bioreceptors. As a result, this aptasensor has been demonstrated to specifically detect thrombin concentrations ranging from 270 pmol/L to 27 nmol/L with an estimated detection limit of 33.5 pmol/L and a response time of ~2 min. Furthermore, it has also been demonstrated that this aptasensor is able to continuously respond to consecutive increasing concentrations of thrombin and to detect binding events as they occur. All these features make this aptasensor a good candidate to continuously study how thrombin concentration progressively increases during the initiation phase of the coagulation cascade.

A Tetra-Panel of Serum Circulating miRNAs for the Diagnosis of the Four Most Prevalent Tumor Types.

The purpose of this study is to clinically validate a series of circulating miRNAs that distinguish between the 4 most prevalent tumor types (lung cancer (LC); breast cancer (BC); colorectal cancer (CRC); and prostate cancer (PCa)) and healthy donors (HDs). A total of 18 miRNAs and 3 housekeeping miRNA genes were evaluated by qRT-PCR on RNA extracted from serum of cancer patients, 44 LC, 45 BC, 27 CRC, and 40 PCa, and on 45 HDs. The cancer detection performance of the miRNA expression levels was evaluated by studying the area under the curve (AUC) of receiver operating characteristic (ROC) curves at univariate and multivariate levels. miR-21 was significantly overexpressed in all cancer types compared with HDs, with accuracy of 67.5% (p = 0.001) for all 4 tumor types and of 80.8% (p < 0.0001) when PCa cases were removed from the analysis. For each tumor type, a panel of miRNAs was defined that provided cancer-detection accuracies of 91%, 94%, 89%, and 77%, respectively. In conclusion, we have described a series of circulating miRNAs that define different tumor types with a very high diagnostic performance. These panels of miRNAs would constitute the basis of different approaches of cancer-detection systems for which clinical utility should be validated in prospective cohorts.

Stabilization of Polymeric Nanofibers Layers for Use as Real-Time and In-Flow Photonic Sensors.

In order to increase the sensitivity of a sensor, the relationship between its volume and the surface available to be functionalized is of great importance. Accordingly, porous materials are becoming very relevant, because they have a notable surface-to-volume ratio. Moreover, they offer the possibility to infiltrate the target substances on them. Among other porous structures, polymeric nanofibers (NFs) layers fabricated by electrospinning have emerged as a very promising alternative to low-cost and easy-to-produce high-performance photonic sensors. However, experimental results show a spectrum drift when performing sensing measurements in real-time. That drift is responsible for a significant error when trying to determine the refractive index variation for a target solution, and, because of that, for the detection of the presence of certain analytes. In order to avoid that problem, different chemical and thermal treatments were studied. The best results were obtained for thermal steps at 190 °C during times between 3 and 5 h. As a result, spectrum drifts lower than 5 pm/min and sensitivities of 518 nm/refractive index unit (RIU) in the visible range of the spectrum were achieved in different electrospun NFs sensors.

Experimental study of an evanescent-field biosensor based on 1D photonic bandgap structures.

A photonic bandgap (PBG) biosensor has been developed for the label-free detection of proteins. As the sensing in this type of structures is governed by the interaction between the evanescent field going into the cladding and the target analytes, scanning near-field optical microscopy has been used to characterize the profile of that evanescent field. The study confirms the strong exponential decrease of the signal as it goes into the cladding. This means that biorecognition events must occur as close to the PBG structure surface as possible in order to obtain the maximum sensing response. Within this context, the PBG biosensor has been biofunctionalized with half-antibodies specific to bovine serum albumin (BSA) using a UV-induced immobilization procedure. The use of half-antibodies allows one to reduce the thickness of the biorecognition volume down to ca. 2.5 nm, thus leading to a higher interaction with the evanescent field, as well as a proper orientation of their binding sites towards the target sample. Then, the biofunctionalized PBG biosensor has been used to perform a direct and real-time detection of the target BSA antigen.

Computational binding study of cardiac troponin I antibody towards cardiac versus skeletal troponin I.

A computational study of the interaction of cardiac troponin I (cTnI) with its specific antibody and of that antibody with skeletal troponin I (sTnI), the principal interferon of cTnI, is carried out. Computational and simulation tools such as FTSite, FTMap, FTDock and pyDock are used to determine the binding sites of these molecules and to study their interactions and molecular docking performance, allowing us to obtain relevant information related with the antigen-antibody interaction for each of the targets. In the context of the development of immunosensing platforms, this type of computational analysis allows performing a preliminary in-silico affinity study of the available bioreceptors for a better selection when moving to the experimental stage, with the subsequent saving in cost and time.

Commercial polycarbonate track-etched membranes as substrates for low-cost optical sensors.

Porous materials have become one of the best options for the development of optical sensors, since they maximize the interaction between the optical field and the target substances, which boosts the sensitivity. In this work, we propose the use of a readily available mesoporous material for the development of such sensors: commercial polycarbonate track-etched membranes. In order to demonstrate their utility for this purpose, we firstly characterized their optical response in the near-infrared range. This response is an interference fringe pattern, characteristic of a Fabry-Pérot interferometer, which is an optical device typically used for sensing purposes. Afterwards, several refractive index sensing experiments were performed by placing different concentrations of ethanol solution on the polycarbonate track-etched membranes. As a result, a sensitivity value of around 56 nm/RIU was obtained and the reusability of the substrate was demonstrated. These results pave the way for the development of optical porous sensors with such easily available mesoporous material.

Thiol-click photochemistry for surface functionalization applied to optical biosensing.

In the field of biosensing, suitable procedures for efficient probes immobilization are of outmost importance. Here we present different light-based strategies to promote the covalent attachment of thiolated capture probes (oligonucleotides and proteins) on different materials and working formats. One strategy employs epoxylated surfaces and uses the light to accomplish the ring opening by a thiol moiety present in a probe. However, most of this work lies on the use of thiol-ene photocoupling chemistry to covalently attach probes to the supports. And thus, both alkenyl and thiol derivatized surfaces are assayed to immobilize thiol or alkene ended probes, respectively, and their performances are compared. Also, the effect of the number of thiols carried by the probe is analyzed comparing single-point and multi-point attachment. The performance of the analogous tethering, but onto alkynylated surfaces is also carried out, and the sensing response is related to the surfaces hydrophobicity. A newly developed reaction is also discussed where a fluorinated surface undergoes the covalent immobilization of thiolated probes activated by light, creating small hydrophilic areas where the probes are attached, and leaving the rest of the surface highly hydrophobic and repellent against protein unspecific adsorption. These mixed surfaces confine the sample (aqueous) uniquely on the hydrophilic spots lowering the background signal and thus increasing the sensitivity. These probe immobilization approaches are applied to fluorescence microarray and label-free nanophotonic biosensing. All the exposed reactions have in common the photoactivation of the thiol moieties, and give rise to quick, clean, versatile, orthogonal and biocompatible reactions. Water is the only solvent used, and light the only catalyzer applied. Thus, all of them can be considered as having the attributes of click-chemistry reactions. For these reasons we named them as thiol-click photochemistry, being a very interesting pool of possibilities when building a biosensor.

Detection of miRNA cancer biomarkers using light activated Molecular Beacons.

Early detection of cancer biomarkers can reduce cancer mortality rate. miRNAs are small non-coding RNAs whose expression changes upon the onset of various types of cancer. Biosensors that specifically detect such biomarkers can be engineered and integrated into point-of-care devices (POC) using label-free detection, high sensibility and compactness. In this paper, a new engineered Molecular Beacon (MB) construct used to detect miRNAs is presented. Such a construct is immobilized onto biosensor surfaces in a covalent and spatially oriented way using the photonic technology Light Assisted Molecular Immobilization (LAMI). The construct consists of a Cy3 labelled MB covalently attached to a light-switchable peptide. One MB construct contains a poly-A sequence in its loop region while the other contains a sequence complementary to the cancer biomarker miRNA-21. The constructs have been characterized by UV-Vis spectroscopy, mass spectrometry and HPLC. LAMI led to the successful immobilization of the engineered constructs onto thiol functionalized optically flat quartz slides and Silicon on Insulator (SOI) sensor surfaces. The immobilized Cy3 labelled MB construct has been imaged using confocal fluorescence microscopy (CFM). The bioavailability of the immobilized engineered MB biosensors was confirmed through specific hybridization with the Cy5 labelled complementary sequence and imaged by CFM and FRET. Hybridization kinetics have been monitored using steady state fluorescence spectroscopy. The label-free detection of miRNA-21 was also achieved by using integrated photonic sensing structures. The engineered light sensitive constructs can be immobilized onto thiol reactive surfaces and are currently being integrated in a POC device for the detection of cancer biomarkers.

Thiol-ene click chemistry towards easy microarraying of half-antibodies.

A UV light-induced thiol-ene coupling reaction (TEC) between half-antibodies (hIgG) and vinyl functionalized glass surfaces was run for biosensing in the microarray format. The accomplished performance improved that obtained with whole antibodies.

High sensitivity and label-free oligonucleotides detection using photonic bandgap sensing structures biofunctionalized with molecular beacon probes.

A label-free sensor, based on the combination of silicon photonic bandgap (PBG) structures with immobilized molecular beacon (MB) probes, is experimentally developed. Complementary target oligonucleotides are specifically recognized through hybridization with the MB probes on the surface of the sensing structure. This combination of PBG sensing structures and MB probes demonstrates an extremely high sensitivity without the need for complex PCR-based amplification or labelling methods.

Experimental study of the evanescent-wave photonic sensors response in presence of molecular beacon conformational changes.

An experimental study of the influence of the conformational change suffered by molecular beacon (MB) probes-upon the biorecognition of nucleic acid target oligonucleotides over evanescent wave photonic sensors-is reported. To this end, high sensitivity photonic sensors based on silicon photonic bandgap (PBG) structures were used, where the MB probes were immobilized via their 5' termination. Those MBs incorporate a biotin moiety close to their 3' termination in order to selectively bind a streptavidin molecule to them. The different photonic sensing responses obtained toward the target oligonucleotide detection, when the streptavidin molecule was bound to the MB probes or not, demonstrate the conformational change suffered by the MB upon hybridization, which promotes the displacement of the streptavidin molecule away from the surface of the photonic sensing structure.

Experimental Study of the Oriented Immobilization of Antibodies on Photonic Sensing Structures by Using Protein A as an Intermediate Layer.

A proper antibody immobilization on a biosensor is a crucial step in order to obtain a high sensitivity to be able to detect low target analyte concentrations. In this paper, we present an experimental study of the immobilization process of antibodies as bioreceptors on a photonic ring resonator sensor. A protein A intermediate layer was created on the sensor surface in order to obtain an oriented immobilization of the antibodies, which enhances the interaction with the target antigens to be detected. The anti-bovine serum albumin (antiBSA)-bovine serum albumin (BSA) pair was used as a model for our study. An opto-fluidic setup was developed in order to flow the different reagents and, simultaneously, to monitor in real-time the spectral response of the photonic sensing structure. The antiBSA immobilization and the BSA detection, their repeatability, and specificity were studied in different conditions of the sensor surface. Finally, an experimental limit of detection for BSA recognition of only 1 ng/mL was obtained.

Real Time Monitoring of a UV Light-Assisted Biofunctionalization Protocol Using a Nanophotonic Biosensor.

A protocol for the covalent biofunctionalization of silicon-based biosensors using a UV light-induced thiol⁻ene coupling (TEC) reaction has been developed. This biofunctionalization approach has been used to immobilize half antibodies (hIgG), which have been obtained by means of a tris(2-carboxyethyl)phosphine (TCEP) reduction at the hinge region, to the surface of a vinyl-activated silicon-on-insulator (SOI) nanophotonic sensing chip. The response of the sensing structures within the nanophotonic chip was monitored in real time during the biofunctionalization process, which has allowed us to confirm that the bioconjugation of the thiol-terminated bioreceptors onto the vinyl-activated sensing surface is only initiated upon UV light photocatalysis.

Real-Time and In-Flow Sensing Using a High Sensitivity Porous Silicon Microcavity-Based Sensor.

Porous silicon seems to be an appropriate material platform for the development of high-sensitivity and low-cost optical sensors, as their porous nature increases the interaction with the target substances, and their fabrication process is very simple and inexpensive. In this paper, we present the experimental development of a porous silicon microcavity sensor and its use for real-time in-flow sensing application. A high-sensitivity configuration was designed and then fabricated, by electrochemically etching a silicon wafer. Refractive index sensing experiments were realized by flowing several dilutions with decreasing refractive indices, and measuring the spectral shift in real-time. The porous silicon microcavity sensor showed a very linear response over a wide refractive index range, with a sensitivity around 1000 nm/refractive index unit (RIU), which allowed us to directly detect refractive index variations in the 10-7 RIU range.

Experimental study of the sensitivity of a porous silicon ring resonator sensor using continuous in-flow measurements.

A highly sensitive photonic sensor based on a porous silicon ring resonator was developed and experimentally characterized. The photonic sensing structure was fabricated by exploiting a porous silicon double layer, where the top layer of a low porosity was used to form photonic elements by e-beam lithography and the bottom layer of a high porosity was used to confine light in the vertical direction. The sensing performance of the ring resonator sensor based on porous silicon was compared for the different resonances within the analyzed wavelength range both for transverse-electric and transverse-magnetic polarizations. We determined that a sensitivity up to 439 nm/RIU for low refractive index changes can be achieved depending on the optical field distribution given by each resonance/polarization.