Label- and Reagent-Free Optical Sensor for Absorption-Based Detection of Urea Concentration in Water Solutions.

Contactless and label-free detection of urea content in aqueous solutions is of great interest in chemical, biomedical, industrial, and automotive applications. In this work, we demonstrate a compact and low-cost instrumental configuration for label-free, reagent-free, and contactless detection of urea dissolved in water, which exploits the absorption properties of urea in the near-infrared wavelength region. The intensity of the radiation transmitted through the fluid under test, contained in a rectangle hollow glass tubing with an optical pathlength of 1 mm, is detected in two spectral bands. Two low-cost, low-power LEDs with emission spectra centered at λ = 1450 nm and λ = 2350 nm are used as readout sources. The photodetector is positioned on the other side of the tubing, in front of the LEDs. The detection performances of a photodiode and of a thermal optical power detector have been compared, exploiting different approaches for LED driving current modulation and photodetected signal processing. The implemented detection system has been tested on urea-water solutions with urea concentrations from 0 up to 525 mg/mL as well as on two samples of commercial diesel exhaust fluid ("AdBlue™"). Considering the transmitted intensity in presence of the urea-water solution, at λ = 1450 nm and λ = 2350 nm, normalized to the transmitted intensity in presence of water, we demonstrate that their ratio is linearly related to urea concentration on a wide range and with good sensitivity.

Optical Multi-Parameter Measuring System for Fluid and Air Bubble Recognition.

Detection of air bubbles in fluidic channels plays a fundamental role in all that medical equipment where liquids flow inside patients' blood vessels or bodies. In this work, we propose a multi-parameter sensing system for simultaneous recognition of the fluid, on the basis of its refractive index and of the air bubble transit. The selected optofluidic platform has been designed and studied to be integrated into automatic pumps for the administration of commercial liquid. The sensor includes a laser beam that crosses twice a plastic cuvette, provided with a back mirror, and a position-sensitive detector. The identification of fluids is carried out by measuring the displacement of the output beam on the detector active surface and the detection of single air bubbles can be performed with the same instrumental scheme, exploiting a specific signal analysis. When a bubble, traveling along the cuvette, crosses the readout light beam, radiation is strongly scattered and a characteristic fingerprint shape of the photo-detected signals versus time is clearly observed. Experimental testing proves that air bubbles can be successfully detected and counted. Their traveling speed can be estimated while simultaneously monitoring the refractive index of the fluid.

Optical Identification of Parenteral Nutrition Solutions Exploiting Refractive Index Sensing.

Parenteral artificial nutrition (PAN) is a lifesaving treatment for a large population of patients affected by different diseases, and it consists of intravenous injection of nutritive fluids by means of infusion pumps. Wrong PAN solutions are, unfortunately, often administered, thus threatening the patients' well-being. Here, we report an optofluidic label-free sensor that can distinguish PAN solutions on the basis of their volumetric refractive index (RI). In our system, a monochromatic light beam, generated by a laser diode, travels obliquely through a transparent, square-section polystyrene channel, is then back-reflected by a mirror, and finally exits the channel in a position that depends on the filling fluid RI. The displacement of the output light spot ΔXexperim is easily detected with a linear, 1-D position sensitive detector (PSD). We initially calibrated the sensor with water-glucose solutions demonstrating a sensitivity S = ΔXexperim/Δn = 13,960 µm/RIU. We then clearly distinguished six commercial PAN solutions, commonly administered to patients. To the best of our knowledge, this is the first reported healthcare sensing platform for remote contactless recognition of PAN fluids, which could be inserted into infusion pumps to improve treatment safety, by checking the compliance to the prescription of the fluid actually delivered to the patient.

Spectral Fingerprint Investigation in the near Infra-Red to Distinguish Harmful Ethylene Glycol from Isopropanol in a Microchannel.

Ethylene glycol (EG) and isopropanol (ISO) are among the major toxic alcohols that pose a risk to human health. However, it is important to distinguish them, since EG is more prone to cause renal failure, and can thus be more dangerous when ingested than ISO. Analysis of alcohols such as isopropanol and ethylene glycol generally can be performed with a complex chromatographic method. Here, we present an optical method based on absorption spectroscopy, performed remotely on EG-ISO mixtures filling a microchannel. Mixtures of ethylene glycol in isopropanol at different volume concentrations were analyzed in a contactless manner in a rectangular-section glass micro-capillary provided with integrated reflectors. Fiber-coupled broadband light in the wavelength range 1.3-1.7 µm crossed the microchannel multiple times before being directed towards an optical spectrum analyzer. The induced zig-zag path increased the fluid-light interaction length and enhanced the effect of optical absorption. A sophisticated theoretical model was developed and the results of our simulations were in very good agreement with the results of the experimental spectral measurements. Moreover, from the acquired data, we retrieved a responsivity parameter, defined as power ratio at two wavelengths, that is linearly related to the EG concentration in the alcoholic mixtures.

Spectral Phase Shift Interferometry for Refractive Index Monitoring in Micro-Capillaries.

In this work, we demonstrate spectral phase-shift interferometry operating in the near-infrared wavelength range for refractive index (RI) monitoring of fluidic samples in micro-capillaries. A detailed theoretical model was developed to calculate the phase-sensitive spectral reflectivity when low-cost rectangular glass micro-capillaries, filled with samples with different refractive indices, are placed at the end of the measurment arm of a Michelson interferometer. From the phase-sensitive spectral reflectivity, we recovered the cosine-shaped interferometric signal as a function of the wavelength, as well as its dependence on the sample RI. Using the readout radiation provided by a 40-nm wideband light source with a flat emission spectrum centered at 1.55 µm and a 2 × 1 fiberoptic coupler on the common input-output optical path, experimental results were found to be in good agreement with the expected theoretical behavior. The shift of the micro-capillary optical resonances, induced by RI variations in the filling fluids (comparing saline solution with respect to distilled water, and isopropanol with respect to ethanol) were clearly detected by monitoring the positions of steep phase jumps in the cosine-shaped interferometric signal recorded as a function of the wavelength. By adding a few optical components to the instrumental configuration previously demonstrated for the spectral amplitude detection of resonances, we achieved phase-sensitive detection of the wavelength positions of the resonances as a function of the filling fluid RI. The main advantage consists of recovering RI variations by detecting the wavelength shift of "sharp peaks", with any amplitude above a threshold in the interferometric signal derivative, instead of "wide minima" in the reflected power spectra, which are more easily affected by uncertainties due to amplitude fluctuations.

Micro-opto-fluidic platform for solvents identification based on absorption properties in the NIR region.

Spectral detection of light transmission through capillaries filled with a fluid sample is a powerful solution for evaluating its composition. In this work, we present an optical method to distinguish water and alcohol samples in a rectangular glass micro-capillary, coupled to an external fluidic path and laid flat onto an aluminum bulk mirror, from the spectral transmittance in the near-infra-red (NIR) wavelength range 1.15-1.65 µm, which becomes sample-specific thanks to the contribution given by the spectral absorption properties of the fluid. The readout beam of broadband radiation is shone on the upper flat side of the micro-capillary with an incidence angle of 14°, crosses the glass walls and the channel depth twice, since it is reflected by the mirror, and it is then coupled to the monochromator input of an optical spectrum analyzer. The theoretical transmission spectra of the capillary filled just with air as well as with distilled water, isopropanol, ethylene glycol, and 95% ethanol (with 5% water content) are derived using analytical equations including the wavelength-dependent attenuation due to fluid absorption. Experimental results relative to the wavelength dependence of the ratio between the spectral transmittance in the presence of the fluid sample and of just air are found to be in agreement with the calculated theoretical behavior. Grapical abstract.

A VCSEL-Based NIR Transillumination System for Morpho-Functional Imaging.

Transillumination with non-ionizing radiation followed by the observation of transmitted and diffused light is the simplest, and probably the oldest method to obtain qualitative information on the internal structure of tissues or body sections. Although scattering precludes formation of high-definition image (unless complex techniques are employed), low resolution pictures complemented by information on the functional condition of the living sample can be extracted. In this context, we have investigated a portable optoelectronic instrumental configuration for efficient transillumination and image detection, even in ambient day-light, of in vivo samples with thickness up to 5 cm, sufficient for visualizing macroscopic structures. Tissue illumination is obtained with an extended source consisting in a matrix of 36 near infrared Vertical Cavity Surface Emitting Lasers (VCSELs) that is powered by a custom designed low-voltage current driver. In addition to the successful acquisition of morphological images of the hand dorsal vein pattern, functional detection of physiological parameters (breath and hearth rate) is achieved non-invasively by means of a monochrome camera, with a Complementary Metal Oxide Semiconductor (CMOS) sensor, turned into a wavelength selective image detector using narrow-band optical filtering.

Characterization of Tunable Micro-Lenses with a Versatile Optical Measuring System.

In this work, we present the results of the opto⁻electro⁻mechanical characterization of tunable micro-lenses, Tlens®, performed with a single-spot optical measuring system. Tested devices are composed of a transparent soft polymer layer that is deposited on a supporting glass substrate and is covered by a glass membrane with a thin-film piezoelectric actuator on top. Near-infrared optical low-coherence reflectometry is exploited for both static and low-frequency dynamic analyses in the time domain. Optical thickness of the layers and of the overall structure, actuation efficiency, and hysteretic behavior of the piezo-actuator as a function of driving voltage are obtained by processing the back-reflected signal in different ways. The use of optical sources with relatively short coherence lengths allows performing interferometric measurements without spurious resonance effects due to multiple parallel interfaces, furthermore, selecting the plane/layer to be monitored. We finally report results of direct measurements of Tlens® optical power as a function of driving voltage, performed by redirecting a He-Ne laser beam on the lens and monitoring the focused spot at various distances with a digital camera.

Near-Infrared Silicon Photonic Crystals with High-Order Photonic Bandgaps for High-Sensitivity Chemical Analysis of Water-Ethanol Mixtures.

Aqueous solutions of alcohols are used in several applications, from pharmaceutics and biology, to chemical, biofuel, and food industries. Nonetheless, development of a simple, inexpensive, and portable sensing device for the quantification of water in water-ethanol mixtures remains a significant challenge. Photonic crystals (PhCs) operating at very high-order photonic bandgaps (PBGs) offer remarkable opportunities for the realization of chemical sensors with high sensitivity and low detection limit. However, high-order PhC structures have been mostly confined to mere theoretical speculations so far, their effective realization requiring microfabrication tools enabling the control of periodic refractive index modulations at the submicrometric scale with extremely high accuracy and precision. Here, we report both experimental and theoretical results on high-sensitivity chemical analysis using vertical, silicon/air 1D-PhCs with spatial period of 10 and 20 µm (namely, over 10 times the operation wavelength) featuring ultra-high-order PBGs in the near-infrared region (namely, up to 50th at 1.1 µm). Fabrication of high-order 1D-PhCs was carried out by electrochemical micromachining (ECM) of silicon, which allowed both surface roughness and deviation from vertical of etched structures to be controlled below 5 nm and 0.1%, respectively. Optical characterization of ECM-fabricated 1D-PhCs, which was performed by acquiring reflectivity spectra over the wavelength range 1-1.7 µm, highlighted the presence of ultra-high-order PBGs with minor optical losses (i.e., <1 dB in reflectivity) separated by deep reflectivity notches with high Q-factors (i.e., >6000), in good agreement with theoretical calculations. Remarkably, the use of high-order 1D-PhCs as refractometric transducers for the quantitative detection of traces of water in water-ethanol mixtures, allowed high sensitivity (namely, either 1000 nm/RIU or ∼0.4 nm/% of water), good detection limit (namely, 5 × 10-3 RIU or ∼10% water), and excellent resolution (namely, either 6 × 10-4 RIU or 1.6% of water) to be reliably achieved on a detection volume of about 168 fL.

Spectral Optical Readout of Rectangular-Miniature Hollow Glass Tubing for Refractive Index Sensing.

For answering the growing demand of innovative micro-fluidic devices able to measure the refractive index of samples in extremely low volumes, this paper presents an overview of the performances of a micro-opto-fluidic sensing platform that employs rectangular, miniature hollow glass tubings. The operating principle is described by showing the analytical model of the tubing, obtained as superposition of different optical cavities, and the optical readout method based on spectral reflectivity detection. We have analyzed, in particular, the theoretical and experimental optical features of rectangular tubings with asymmetrical geometry, thus with channel depth larger than the thickness of the glass walls, though all of them in the range of a few tens of micrometers. The origins of the complex line-shape of the spectral response in reflection, due to the different cavities formed by the tubing flat walls and channel, have been investigated using a Fourier transform analysis. The implemented instrumental configuration, based on standard telecom fiberoptic components and a semiconductor broadband optical source emitting in the near infrared wavelength region centered at 1.55 µm, has allowed acquisition of reflectivity spectra for experimental verification of the expected theoretical behavior. We have achieved detection of refractive index variations related to the change of concentration of glucose-water solutions flowing through the tubing by monitoring the spectral shift of the optical resonances.

Flow-through micro-capillary refractive index sensor based on T/R spectral shift monitoring.

We present a flow-through refractive index sensor for measuring the concentration of glucose solutions based on the application of rectangular glass micro-capillaries, with channel depth of 50 µm and 30 µm. A custom designed and 3D printed polymeric shell protects the tiny capillaries, ensuring an easier handling and interconnection with the macro-fluidic path. By illuminating the capillary with broadband radiation centered at λ~1.55 µm, both the transmitted (T) and reflected (R) optical spectrum from the capillary are detected with an optical spectrum analyzer, exploiting an all-fiber setup. Monitoring the spectral shift of the ratio T/R in response to increasing concentration of glucose solutions in water we have obtained sensitivities up to 530.9 nm/RIU and limit of detection in the range of 10-5-10-4 RIU. Experimental results are in agreement with the theoretically predicted principle of operation. After the demonstration of amplitude detection at a single wavelength, we finally discuss the impact of the capillary parameters on the sensitivity.

Testing of Piezo-Actuated Glass Micro-Membranes by Optical Low-Coherence Reflectometry.

In this work, we have applied optical low-coherence reflectometry (OLCR), implemented with infra-red light propagating in fiberoptic paths, to perform static and dynamic analyses on piezo-actuated glass micro-membranes. The actuator was fabricated by means of thin-film piezoelectric MEMS technology and was employed for modifying the micro-membrane curvature, in view of its application in micro-optic devices, such as variable focus micro-lenses. We are here showing that OLCR incorporating a near-infrared superluminescent light emitting diode as the read-out source is suitable for measuring various parameters such as the micro-membrane optical path-length, the membrane displacement as a function of the applied voltage (yielding the piezo-actuator hysteresis) as well as the resonance curve of the fundamental vibration mode. The use of an optical source with short coherence-time allows performing interferometric measurements without spurious resonance effects due to multiple parallel interfaces of highly planar slabs, furthermore selecting the plane/layer to be monitored. We demonstrate that the same compact and flexible setup can be successfully employed to perform spot optical measurements for static and dynamic characterization of piezo-MEMS in real time.

Low-Coherence Reflectometry for Refractive Index Measurements of Cells in Micro-Capillaries.

The refractive index of cells provides insights into their composition, organization and function. Moreover, a good knowledge of the cell refractive index would allow an improvement of optical cytometric and diagnostic systems. Although interferometric techniques undoubtedly represent a good solution for quantifying optical path variation, obtaining the refractive index of a population of cells non-invasively remains challenging because of the variability in the geometrical thickness of the sample. In this paper, we demonstrate the use of infrared low-coherence reflectometry for non-invasively quantifying the average refractive index of cell populations gently confined in rectangular glass micro-capillaries. A suspension of human red blood cells in plasma is tested as a reference. As a use example, we apply this technique to estimate the average refractive index of cell populations belonging to epithelial and hematological families.

3D Silicon Microstructures: A New Tool for Evaluating Biological Aggressiveness of Tumor Cells.

In this work, silicon micromachined structures (SMS), consisting of arrays of 3- µ m-thick silicon walls separated by 50- µm-deep, 5- µ m-wide gaps, were applied to investigate the behavior of eight tumor cell lines, with different origins and biological aggressiveness, in a three-dimensional (3D) microenvironment. Several cell culture experiments were performed on 3D-SMS and cells grown on silicon were stained for fluorescence microscopy analyses. Most of the tumor cell lines recognized in the literature as highly aggressive (OVCAR-5, A375, MDA-MB-231, and RPMI-7951) exhibited a great ability to enter and colonize the narrow deep gaps of the SMS, whereas less aggressive cell lines (OVCAR-3, Capan-1, MCF7, and NCI-H2126) demonstrated less penetration capability and tended to remain on top of the SMS. Quantitative image analyses of several fluorescence microscopy fields of silicon samples were performed for automatic cell recognition and count, in order to quantify the fraction of cells inside the gaps, with respect to the total number of cells in the examined field. Our results show that higher fractions of cells in the gaps are obtained with more aggressive cell lines, thus supporting in a quantitative way the observation that the behavior of tumor cells on the 3D-SMS depends on their aggressiveness level.

Label-free optical detection of cells grown in 3D silicon microstructures.

We demonstrate high aspect-ratio photonic crystals that could serve as three-dimensional (3D) microincubators for cell culture and also provide label-free optical detection of the cells. The investigated microstructures, fabricated by electrochemical micromachining of standard silicon wafers, consist of periodic arrays of silicon walls separated by narrow deeply etched air-gaps (50 µm high and 5 µm wide) and feature the typical spectral properties of photonic crystals in the wavelength range 1.0-1.7 µm: their spectral reflectivity is characterized by wavelength regions where reflectivity is high (photonic bandgaps), separated by narrow wavelength regions where reflectivity is very low. In this work, we show that the presence of cells, grown inside the gaps, strongly affects light propagation across the photonic crystal and, therefore, its spectral reflectivity. Exploiting a label-free optical detection method, based on a fiberoptic setup, we are able to probe the extension of cells adherent to the vertical silicon walls with a non-invasive direct testing. In particular, the intensity ratio at two wavelengths is the experimental parameter that can be well correlated to the cell spreading on the silicon wall inside the gaps.

Isolation of Langerhans islets by dielectrophoresis.

The purification of Langerhans islets from fragments of pancreatic exocrine tissue is a critical stage for the further transplantation of insulin secreting islets in patients affected by type I diabetes. Aim of our work was the evaluation of dielectrophoresis as a promising method for pancreatic islets isolation without physical contact in miniaturized lab-on-chip devices. DEP exploits the dielectric properties of particles suspended in a fluid, in a region where the amplitude of the electric field is characterized by a high gradient. Langerhans islets are aggregates of cells and have a minimum diameter of 50 microns. Dielectric models of pancreatic islets as cell aggregates were derived from single pancreatic beta cells model. Numerical simulations were performed to optimize the exact shape and size of the quadrupole microelectrode configuration and to determine the DEP forces acting on islets. A custom electronic setup was developed for the generation of sinusoidal signals with proper voltage and frequency and used to perform DEP experiments with samples of Langerhans islets. Dielectric models were found sufficiently accurate and negative DEP, showing repulsion from the electrodes, was observed for pancreatic islets. The results of this work demonstrate that Langerhans islet can be manipulated without physical contact by dielectrophoresis, a technique that can be applied on cell aggregates in miniaturized lab-on-chip devices.

A new cell-selective three-dimensional microincubator based on silicon photonic crystals.

In this work, we show that vertical, high aspect-ratio (HAR) photonic crystals (PhCs), consisting of periodic arrays of 5 µm wide gaps with depth of 50 µm separated by 3 µm thick silicon walls, fabricated by electrochemical micromachining, can be used as three-dimensional microincubators, allowing cell lines to be selectively grown into the gaps. Silicon micromachined dice incorporating regions with different surface profiles, namely flat silicon and deeply etched PhC, were used as microincubators for culturing adherent cell lines with different morphology and adhesion properties. We extensively investigated and compared the proliferative behavior on HAR PhCs of eight human cell models, with different origins, such as the epithelial (SW613-B3; HeLa; SW480; HCT116; HT29) and the mesenchymal (MRC-5V1; CF; HT1080). We also verified the contribution of cell sedimentation into the silicon gaps. Fluorescence microscopy analysis highlights that only cell lines that exhibit, in the tested culture condition, the behavior typical of the mesenchymal phenotype are able to penetrate into the gaps of the PhC, extending their body deeply in the narrow gaps between adjacent silicon walls, and to grow adherent to the vertical surfaces of silicon. Results reported in this work, confirmed in various experiments, strongly support our statement that such three-dimensional microstructures have selection capabilities with regard to the cell lines that can actively populate the narrow gaps. Cells with a mesenchymal phenotype could be exploited in the next future as bioreceptors, in combination with HAR PhC optical transducers, e.g., for label-free optical detection of cellular activities involving changes in cell adhesion and/or morphology (e.g., apoptosis) in a three-dimensional microenvironment.

Fibrillogenesis of human ß2 -microglobulin in three-dimensional silicon microstructures.

The authors describe the interaction of biological nanostructures formed by ß(2) -microglobulin amyloid fibrils with three-dimensional silicon microstructures consisting in periodic arrays of vertical silicon walls (≈3 µm-thick) separated by 50 µm-deep air gaps (≈5 µm-wide). These structures are of great interest from a biological point of view since they well mimic the interstitial environment typical of amyloid deposition in vivo. Moreover, they behave as hybrid photonic crystals, potentially applicable as optical transducers for label-free detection of the kinetics of amyloid fibrils formation. Fluorescence and atomic force microscopy (AFM) show that a uniform distribution of amyloid fibrils is achieved when fibrillogenesis occurs directly on silicon. The high resolution AFM images also demonstrate that amyloid fibrils grown on silicon are characterized by the same fine structure typically ensured by fibrillogenesis in solution.

Optical characterization of alcohol-infiltrated one-dimensional silicon photonic crystals.

In this work, experimental results on the optical characterization of alcohol-infiltrated silicon/air one-dimensional photonic crystals (1D-PhCs), fabricated by electrochemical micromachining of silicon, are presented. The spectral reflectivity of high-order hybrid 1D-PhCs with a spatial period of 8 microm was measured, in the wavelength range 1.0-1.7 microm, when alcohols (ethanol and isopropanol) substitute air inside the trenches. A reliable redshift is observed in the presence of alcohols, with respect to air, which allows one to discriminate the refractive index difference between the alcohols. Experimental data are in good agreement with numerical results calculated by using the characteristic matrix method, modified to take into account surface roughness of silicon walls.