Photonic Dipstick Immunosensor to Detect Adulteration of Ewe, Goat, and Donkey Milk with Cow Milk through Bovine κ-Casein Detection.

The quality and authenticity of milk are of paramount importance. Cow milk is more allergenic and less nutritious than ewe, goat, or donkey milk, which are often adulterated with cow milk due to their seasonal availability and higher prices. In this work, a silicon photonic dipstick sensor accommodating two U-shaped Mach-Zehnder Interferometers (MZIs) was employed for the label-free detection of the adulteration of ewe, goat, and donkey milk with cow milk. One of the two MZIs of the chip was modified with bovine κ-casein, while the other was modified with bovine serum albumin to serve as a blank. All assay steps were performed by immersion of the chip side where the MZIs are positioned into the reagent solutions, leading to a photonic dipstick immunosensor. Thus, the chip was first immersed in a mixture of milk with anti-bovine κ-casein antibody and then in a secondary antibody solution for signal enhancement. A limit of detection of 0.05% v/v cow milk in ewe, goat, or donkey milk was achieved in 12 min using a 50-times diluted sample. This fast, sensitive, and simple assay, without the need for sample pre-processing, microfluidics, or pumps, makes the developed sensor ideal for the detection of milk adulteration at the point of need.

Determining the Nutrient Content of Hydroponically-Cultivated Microgreens with Immersible Silicon Photonic Sensors: A Preliminary Feasibility Study.

Microgreens have gained attention for their exceptional culinary characteristics and high nutritional value. The present study focused on a novel approach for investigating the easy extraction of plant samples and the utilization of immersible silicon photonic sensors to determine, on the spot, the nutrient content of microgreens and their optimum time of harvest. For the first time, it was examined how these novel sensors can capture time-shifting spectra caused by the molecules' dynamic adhesion onto the sensor surface. The experiment involved four types of microgreens (three types of basil and broccoli) grown in a do-it-yourself hydroponic installation. The sensors successfully distinguished between different plant types, showcasing their discriminative capabilities. To determine the optimum harvest time, this study compared the sensor data with results obtained through standard analytical methods. Specifically, the total phenolic content and antioxidant activity of two basil varieties were juxtaposed with the sensor data, and this study concluded that the ideal harvest time for basil microgreens was 14 days after planting. This finding highlights the potential of the immersible silicon photonic sensors for potentially replacing time-consuming analytical techniques. By concentrating on obtaining plant extracts, capturing time-shifting spectra, and assessing sensor reusability, this research paves the way for future advancements in urban farming.

Rapid detection of mozzarella and feta cheese adulteration with cow milk through a silicon photonic immunosensor.

Mozzarella di Bufala Campana and Feta are two cheeses with Protected Designation of Origin the fraudulent adulteration of which with bovine milk must be routinely checked to ensure that consumers actually buy these high-end products and avoid health issues related to bovine milk allergy. Here, we employed, for the first time, a silicon-based photonic immunosensor for the detection of mozzarella and feta adulteration with bovine milk. The photonic immunosensor used relies on Mach-Zehnder interferometers monolithically integrated along with their respective light sources on a silicon chip. A rabbit polyclonal antiserum raised against bovine κ-casein was used for the development of a competitive immunoassay realized in three steps, including a reaction with the antiserum, a biotinylated anti-rabbit IgG antibody, and streptavidin. The implementation of this assay configuration significantly reduced the non-specific signal due to the cheese matrix, and allowed completion of the assay in ∼9 min. After optimization of all assay conditions, bovine cheese could be quantified in mozzarella or feta at concentrations as low as 0.5 and 0.25% (w/w), respectively; both quantification limits were below the maximum allowable content of bovine milk in mozzarella and feta (1% w/w) according to the EU regulations. Equally important, the assays were reproducible with intra- and inter-assay coefficients of variation <10%, and exhibited a wide linear dynamic range that extended up to 50 and 25% (w/w) for mozzarella and feta, respectively. Taking into account its performance, the proposed immunosensor may be transformed to a new tool against fraudulent activities in the dairy industry.

Ultrafast Multiplexed-Allergen Detection through Advanced Fluidic Design and Monolithic Interferometric Silicon Chips.

A silicon-based miniaturized sensor chip combined with an advanced microfluidic module for the simultaneous, label-free immunochemical determination of four allergens, bovine milk protein, peanut protein, soy protein, and gliadin, is presented. The sensor chip consists of an array of 10 broad-band Mach-Zehnder interferometers (BB-MZIs) monolithically integrated on silicon, along with their respective broad-band light sources. The BB-MZIs were biofunctionalized with the targeted allergens and their responses during immunoreaction were monitored by multiplexing their transmission spectra through an external miniaturized spectrometer. The assay is performed by running mixtures of calibrators or samples with the antibodies against the four allergens followed by an antispecies specific antibodies solution. Employing a fluidic module of nearly one-dimensional geometry, that provided for uniform delivery of the reagents, CV values <6% were achieved for the responses of the 10 BB-MZIs, allowing for reliable multianalyte determinations. The analysis is completed in 6.5 min, and the detection limits were 0.04 µg/mL for bovine k-casein, 1.0 µg/mL for peanut protein, 0.80 µg/mL for soy protein, and 0.10 µg/mL for gliadin. The assays were accurate (recoveries 88-118%) and repeatable (intra- and interassay CVs <7% for all four allergens). Finally, the sensor was evaluated by analyzing samples from a cleaning in place system (CIP) of a dairy industry and the results obtained were in good agreement with those received by the respective ELISAs. The analytical characteristics of the sensor combined with the short analysis time and the small chip size make the proposed system an ideal tool for on-site multianalyte determinations.

Humidity Sensing Properties of Paper Substrates and Their Passivation with ZnO Nanoparticles for Sensor Applications.

In this paper, we investigated the effect of humidity on paper substrates and propose a simple and low-cost method for their passivation using ZnO nanoparticles. To this end, we built paper-based microdevices based on an interdigitated electrode (IDE) configuration by means of a mask-less laser patterning method on simple commercial printing papers. Initial resistive measurements indicate that a paper substrate with a porous surface can be used as a cost-effective, sensitive and disposable humidity sensor in the 20% to 70% relative humidity (RH) range. Successive spin-coated layers of ZnO nanoparticles then, control the effect of humidity. Using this approach, the sensors become passive to relative humidity changes, paving the way to the development of ZnO-based gas sensors on paper substrates insensitive to humidity.

Assessment of goat milk adulteration with a label-free monolithically integrated optoelectronic biosensor.

The label-free detection of bovine milk in goat milk through a miniaturized optical biosensor is presented. The biosensor consists of ten planar silicon nitride waveguide Broad-Band Mach-Zehnder interferometers (BB-MZIs) monolithically integrated and self-aligned with their respective silicon LEDs on the same Si chip. The BB-MZIs were transformed to biosensing transducers by functionalizing their sensing arm with bovine k-casein. Measurements were performed by continuously recording the transmission spectra of each interferometer through an external spectrometer. The amount of bovine milk in goat milk was determined through a competitive immunoassay by passing over the sensor mixtures of anti-k-casein antibodies with the calibrators or the samples. The output spectra of each BB-MZI recorded during the reaction were subjected to Discrete Fourier Transform in order to convert the observed spectral shifts to phase shifts in the wavenumber domain. The method had a detection limit of 0.04 % (v/v) bovine milk in goat milk, dynamic range 0.1-1.0 % (v/v), recoveries 93-110 %, and intra- and inter-assay coefficients of variation less than 12 and 15 %, respectively. The proposed biosensor compared well in terms of analytical performance with a competitive ELISA developed using the same monoclonal antibodies. Nevertheless, the duration of the biosensor assay was 10 min whereas the ELISA required 2 h. Thus, the fast and sensitive determinations along with the small size of the sensor make it ideal for incorporation into portable devices for assessment of goat or ewe's milk adulteration with bovine milk at the point-of-need.

Directly immersible silicon photonic probes: Application to rapid SARS-CoV-2 serological testing.

Silicon photonic probes based on broad-band Mach-Zehnder interferometry are explored for the first time as directly immersible immunosensors alleviating the need for microfluidics and pumps. Each probe includes two U-shaped waveguides allowing light in- and out-coupling from the same chip side through a bifurcated fiber and a mechanical coupler. At the opposite chip side, two Mach-Zehnder interferometers (MZI) are located enabling real-time monitoring of binding reactions by immersion of this chip side into a sample. The sensing arm windows of the two MZIs have different length resulting in two distinct peaks in the Fourier domain, the phase shift of which can be monitored independently through Fast Fourier Transform of the output spectrum. The photonic probes analytical potential was demonstrated through detection of antibodies against SARS-CoV-2 in human serum samples. For this, one MZI was functionalized with the Receptor Binding Domain (RBD) of SARS-CoV-2 Spike 1 protein, and the other with bovine serum albumin to serve as reference. The biofunctionalized probes were immersed for 10 min in human serum sample and then for 5 min in goat anti-human IgG Fc specific antibody solution. Using a humanized rat antibody against SARS-CoV-2 RBD, a detection limit of 20 ng/mL was determined. Analysis of human serum samples indicated that the proposed sensor discriminated completely non-infected/non-vaccinated from vaccinated individuals, and the antibodies levels determined correlated well with those determined in the same samples by ELISA. These results demonstrated the potential of the proposed sensor to serve as an efficient tool for expeditious point-of-care testing.

CuO/PMMA Polymer Nanocomposites as Novel Resist Materials for E-Beam Lithography.

Polymer nanocomposites have emerged as a new powerful class of materials because of their versatility, adaptability and wide applicability to a variety of fields. In this work, a facile and cost-effective method to develop poly(methyl methacrylate) (PMMA)-based polymer nanocomposites with copper oxide (CuO) nanofillers is presented. The study concentrates on finding an appropriate methodology to realize CuO/PMMA nanocomposites that could be used as resist materials for e-beam lithography (EBL) with the intention of being integrated into nanodevices. The CuO nanofillers were synthesized via a low-cost chemical synthesis, while several loadings, spin coating conditions and two solvents (acetone and methyl ethyl ketone) were explored and assessed with regards to their effect on producing CuO/PMMA nanocomposites. The nanocomposite films were patterned with EBL and contrast curve data and resolution analysis were used to evaluate their performance and suitability as a resist material. Micro-X-ray fluorescence spectroscopy (µ-XRF) complemented with XRF measurements via a handheld instrument (hh-XRF) was additionally employed as an alternative rapid and non-destructive technique in order to investigate the uniform dispersion of the nanofillers within the polymer matrix and to assist in the selection of the optimum preparation conditions. This study revealed that it is possible to produce low-cost CuO/PMMA nanocomposites as a novel resist material without resorting to complicated preparation techniques.

Integrated optical frequency-resolved Mach-Zehnder interferometers for label-free affinity sensing.

Integrated Optical Frequency-Resolved Mach-Zehnder Interferometry (IO FR-MZI) is introduced as an alternative, cost-efficient operation principle for integrated optical label-free affinity sensors that can combine high sensitivity with high versatility in terms of potential applications and experimental configurations. A detailed theoretical analysis of the method is presented followed by a semi-analytical approximation and numerical calculations in order to quantify the sensitivity and limits of detection of the FR-MZI over Single Wavelength MZI. The obtained results substantiate that IO FR-MZI- based sensors constitute a generic technological platform of high sensitivity that can be implemented into a plethora of detection schemes. For an optimized optical design well below 1mm in length the limit of detection can be as low as 0.025A in terms of adlayer effective thickness allowing for truly miniaturized integrated optical sensors fabricated with high yield with standard microfabrication techniques.

Simultaneous determination of aflatoxin B1, fumonisin B1 and deoxynivalenol in beer samples with a label-free monolithically integrated optoelectronic biosensor.

A label-free optical biosensor for the fast simultaneous determination of three mycotoxins, aflatoxin B1 (AFB1), fumonisin B1 (FB1) and deoxynivalenol (DON), in beer samples is presented. The biosensor is based on an array of ten Mach-Zehnder interferometers (MZIs) monolithically integrated along with their respective broad-band silicon light sources onto a single chip. Multi-analyte determination is accomplished by functionalizing the sensing arms of individual MZIs with mycotoxin-protein conjugates. Assay is performed by pumping over the chip mixtures of calibrators or samples with a mixture of specific monoclonal antibodies, followed by reaction with a secondary anti-mouse IgG antibody. Reactions are monitored in real-time by continuously recording the MZI output spectra, which are then subjected to Discrete Fourier Transform to convert spectrum shifts to phase shifts. The detection limits achieved for AFB1, FB1 and DON were 0.8, 5.6 and 24 ng/ml, respectively, while the assay duration was 12 min. Recovery values ranging from 85 to 115% were determined in beer samples spiked with known concentrations of the three mycotoxins. In addition, beers of different types and origin were analysed with the biosensor developed and the results were compared with those provided by established laboratory methods, further supporting the accuracy of the proposed device.

Fast label-free detection of C-reactive protein using broad-band Mach-Zehnder interferometers integrated on silicon chips.

An immunosensor for fast and accurate determination of C-reactive protein (CRP) in human serum samples based on an array of all-silicon broad-band Mach-Zehnder interferometers (BB-MZIs) is demonstrated. The detection was based on monitoring the spectral shifts during the binding of CRP on the antibody molecules that have been immobilized on the sensing arms of the BB-MZIs. By employing the reaction rate as the analytical signal the assay time was compressed to few minutes. The detection limit was 2.1ng/mL, the quantification limit was 4.2ng/mL and the linear dynamic range extended up to 100ng/mL. The measurements performed in human serum samples with the developed immunosensor were characterized by high repeatability and accuracy as it was demonstrated by dilution linearity and recovery experiments. In addition, the concentration values determined were in excellent agreement with those determined for the same samples by a standard clinical laboratory method. The compact size of the chip makes the proposed immunosensor attractive for incorporation into miniaturized devices for the determination of clinical analytes at the point-of-need.

Detection of ochratoxin A in beer samples with a label-free monolithically integrated optoelectronic biosensor.

An optical biosensor for label-free detection of ochratoxin A (OTA) in beer samples is presented. The biosensor consists of an array of ten Mach-Zehnder interferometers (MZIs) monolithically integrated along with their respective broad-band silicon light sources on the same Si chip (37mm2). The chip was transformed to biosensor by functionalizing the MZIs sensing arms with an OTA-ovalbumin conjugate. OTA determination was performed by pumping over the chip mixtures of calibrators or samples with anti-OTA antibody following a competitive immunoassay format. An external miniaturized spectrometer was employed to continuously record the transmission spectra of each interferometer. Spectral shifts obtained due to immunoreaction were transformed to phase shifts through Discrete Fourier Transform. The assay had a detection limit of 2.0ng/ml and a dynamic range 4.0-100ng/ml in beer samples, recoveries ranging from 90.6 to 116%, and intra- and inter-assay coefficients of variation of 9% and 14%, respectively. The results obtained with the sensor using OTA-spiked beer samples spiked were in good agreement with those obtained by an ELISA developed using the same antibody. The good analytical performance of the biosensor and the small size of the proposed chip provide for the development of a portable instrument for point-of-need determinations.

ZnO Nanostructure Templates as a Cost-Efficient Mass-Producible Route for the Development of Cellular Networks.

The development of artificial surfaces which can regulate or trigger specific functions of living cells, and which are capable of inducing in vivo-like cell behaviors under in vitro conditions has been a long-sought goal over the past twenty years. In this work, an alternative, facile and cost-efficient method for mass-producible cellular templates is presented. The proposed methodology consists of a cost-efficient, two-step, all-wet technique capable of producing ZnO-based nanostructures on predefined patterns on a variety of substrates. ZnO-apart from the fact that it is a biocompatible material-was chosen because of its multifunctional nature which has rendered it a versatile material employed in a wide range of applications. Si, Si3N4, emulated microelectrode arrays and conventional glass cover slips were patterned at the micrometer scale and the patterns were filled with ZnO nanostructures. Using HeLa cells, we demonstrated that the fabricated nanotopographical features could promote guided cellular adhesion on the pre-defined micron-scale patterns only through nanomechanical cues without the need for further surface activation or modification. The basic steps of the micro/nanofabrication are presented and the results from the cell adhesion experiments are discussed, showing the potential of the suggested methodology for creating low-cost templates for engineered cellular networks.

Monolithically integrated Mach-Zehnder biosensors for real-time label-free monitoring of biomolecular reactions.

Arrays of monolithically integrated Mach-Zehnder interferometers were fabricated by standard silicon technology and applied to the label-free real-time monitoring of biomolecular interactions. Chips accommodating 10 MZIs were functionalized with recognition biomolecules and encapsulated in wafer scale. Detection is based on Frequency-Resolved Mach-Zehnder Interferometry, a new concept that takes advantage of the broad-band input spectrum by monitoring the changes for every input frequency. The sensitivity of the device in terms of refractive index changes (Δn) was calculated using isopropanol/water solutions. A detection limit of Δn = 4 × 10(-6) was calculated. The bioanalytical capabilities of the device there demonstrated through model binding assays (biotin/streptavidin) as well as the detection of total prostate specific antigen in serum samples using devices coated with antigen-specific monoclonal antibody. Detection limits at the pM range were determined.

Monolithically integrated biosensors based on Frequency-Resolved Mach-Zehnder Interferometers for multi-analyte determinations.

The application of fully monolithically-integrated Mach-Zehnder interferometer arrays fabricated by standard silicon technology to the label-free detection of analytes is introduced. Detection with the presented biosensor is based on a novel concept, the Frequency-Resolved Mach-Zehnder Interferometry (FR-MZI). In addition, a smart encapsulation based on an appropriately designed microfluidic system and performed at the wafer scale scheme for the easy delivery of the samples to be analyzed is demonstrated. Testing of the FR-MZI biosensors with model binding assays demonstrated the detection of streptavidin binding to immobilized biotin at concentrations in the sub nM range. This is the first experimental demonstration of the FR-MZI concept as well as the first demonstration of a monolithically fully-integrated MZI biosensor.

Polyoxometalate-based layered structures for charge transport control in molecular devices.

Hybrid organic-inorganic films consisted of molecular layers of a Keggin-structure polyoxometalate (POM: 12-tungstophosphoric acid, H(3)PW(12)O(40)) and 1,12-diaminododecane (DD) on 3-aminopropyl triethoxysilane (APTES)-modified silicon surface, fabricated via the layer-by-layer (LBL) self-assembly method are evaluated as molecular materials for electronic devices. The effect of the fabrication process parameters, including primarily compositions of deposition solutions, on the structural characteristics of the POM-based multilayers was studied extensively with a combination of spectroscopic methods (UV, FTIR, and XPS). Well-characterized POM-based films (both single-layers and multilayers) in a controlled and reproducible way were obtained. The conduction mechanisms in single-layered and multilayered structures were elucidated by the electrical characterization of the produced films supported by the appropriate theoretical analysis. Fowler-Nordheim (FN) tunneling and percolation mechanisms were encountered in good correlation with the structural characteristics of the films encouraging further investigation on the use of these materials in electronic and, in particular, in memory devices.