Enzymatically mediated fluorescent copper nanocluster generation for tyramine determination.

This work details the enzymatic generation of fluorescence nanomaterials and the use of this optical signal as the analytical parameter for the quantification of the substrate. More specifically, fluorescent copper nanoclusters have been obtained during the enzymatic reaction of tyramine oxidase and tyramine in the presence of Cu(II); the fluorescence intensity being proportional to the concentration of tyramine. The nanoclusters obtained show fluorescence at 445 nm by being excited at 320 nm and have been characterized by TEM, EDX, and XPS. The formation mechanism has also been studied, suggesting that under the optimal conditions (0.1 M MES buffer and pH = 6), the formation of the nanoclusters is due to the reducing properties of the product of the enzymatic reaction (p-hydroxybenzaldehyde) in MES buffer. The method shows a linear relationship with the concentration of tyramine in the range from 1.0·10-5 to 2.5·10-4 M, a RSD of 3% (n = 5) and a LOD of 6.3·10-6 M. The method has been applied to the determination of tyramine in sausage with good results.

Portable colorimetric enzymatic disposable biosensor for histamine and simultaneous histamine/tyramine determination using a smartphone.

Tyramine oxidase (TAO), peroxidase (HRP), and Amplex Red (AR) have been immobilized on cellulose to obtain disposable biosensors for the determination of histamine. During the enzymatic reaction, AR is oxidized and a pink spot is obtained. Using a smartphone and measuring the G (green) color coordinate, histamine can be determined in the presence of other biogenic amines (putrescine and cadaverine) in concentrations ranging from 2·10-5 M to 5·10-4 M with a 7.5·10-6 M limit of detection (LoD). Despite tyramine interference, experimental conditions are provided which allow rapid and simple histamine and simultaneous histamine/tyramine (semi)quantitative determination in mixtures. Finally, tyramine and histamine were determined in a tuna extract with good results (compared to the reference HPLC-MS method). The methodology can also be applied in solution allowing histamine (and simultaneous histamine/tyramine) determination with a lower LoD (1.8·10-7 M) and a similar selectivity.

Poster Session: Experimental assessment of scleral anisotropy using multi-meridian air-coupled ultrasonic optical coherence elastography.

Scleral biomechanics plays a key role in the understanding of myopia progression. In this study, we characterized the elastic properties of sclera using an air-coupled ultrasonic (ACUS) optical coherence elastography (OCE) system. New Zealand rabbit eyes (n=7) were measured (<24hr postmortem) in four scleral locations: superior/inferior temporal (ST, IT), and superior/inferior nasal (SN, IN) maintaining an intraocular pressure of 15 mmHg. Elastic waves were induced in the sclera, and wave propagation velocity and shear modulus were measured along two directions: circumferential (superior-inferior) and meridional (nasal-temporal). Wave velocity in scleral tissue ranged from 6 to 24 m/s and shear modulus from 11 to 150 kPa. Velocity was significantly higher (p<.001) in the circumferential vs. meridional directions in the following locations: ST:15.83±2.85 vs 9.43±1.68 m/s, IT:15.00±3.98 vs 8.93±1.53 m/s; SN:16.79±4.30 vs 9.27±1.47 m/s; and IN:13.92±3.85 vs 8.57±1.46 m/s. The average shear modulus in the circumferential was also significantly higher (p<.001) than in the meridional direction for all locations: 65.37±6.04 vs 22.55±1.36 kPa. These results show that the rabbit sclera is mechanically anisotropic with higher rigidity in the circumferential direction compared to the meridional direction. ACUS-OCE is a promising non-invasive method to quantify the biomechanical changes in scleral tissue for future studies involving myopia treatments.

Poster Session: Estimation of scleral biomechanical properties from air-puff-coupled optical coherence tomography.

Progression of myopia is usually accompanied by axial overgrowth of the eyeball, which affects scleral biomechanics (BM). To study scleral biomechanics, we propose the use of air-puff deformation swept-source OCT imaging. Air-puff deformation imaging was performed at different sites of ex vivo porcine (n=5) and rabbit (n=3) eyes, (<24hr postmortem): Nasal/temporal equatorial and posterior sclera (NE, NP, TE, TP), superior (S) and inferior (I) sclera, and cornea (C). Intraocular pressure was kept at 15mmHg. Deformation data were used as input to inverse finite element model (FEM) algorithms to reconstruct BM properties. Experimental deformation amplitudes showed dependence on the animal model, with porcine scleras exhibiting greater inter-site variation (displacement of S, I was up to four times greater than that of N, T), while rabbit scleras exhibited at most 40% of displacement differences between all sites. Both models showed significant (p<.001) differences in the temporal deformation profile between sclera and (C), but similarities in all scleral locations, suggesting that the scleral temporal profile is independent of scleral thickness variations. The FEM estimated an elastic modulus of 1.84 ± 0.30 MPa (I) to 6.04 ± 2.11 MPa (TE) for the porcine sclera. The use of scleral air-puff imaging is promising for noninvasive investigation of structural changes in the sclera associated with myopia and for monitoring possible modulation of scleral stiffness with myopia treatment.

In situ enzymatic generation of Au/Pt nanoparticles as an analytical photometric system: proof of concept determination of tyramine.

In situ enzymatic generation of bimetallic nanoparticles, mainly Au/Pt, overcomes the drawbacks (continuous absorbance drift, modest LOQ, and long-time reaction) observed when AuNP alone are produced. In this study, Au/Pt nanoparticles have been characterized by EDS, XPS, and HRTEM images using the enzymatic determination of tyramine with tyramine oxidase (TAO) as a model. Under experimental conditions, the Au/Pt NPs show an absorption maximum at 580 nm which can be related to the concentration of tyramine in the range 1.0 × 10-6M to 2.5 × 10-4M with a RSD of 3.4% (n = 5, using 5 × 10-6M tyramine). The Au/Pt system enables low LOQ (1.0 × 10-6 M), high reduction of the absorbance drift, and a significant shortening of the reaction time (i.e., from 30 to 2 min for a [tyramine] = 1 × 10-4M); additionally, a better selectivity is also obtained. The method has been applied to tyramine determination in cured cheese and no significant differences were obtained compared to a reference method (HRP:TMB). The effect of Pt(II) seems to involve the previous reduction of Au(III) to Au(I) and NP generation from this oxidation state. Finally, a three-step (nucleation-growth-aggregation) kinetic model for the generation of NPs is proposed; this has enabled us to obtain a mathematical equation which explains the experimentally observed variation of the absorbance with time.

Tectomer-Mediated Optical Nanosensors for Tyramine Determination.

The development of optical sensors for in situ testing has become of great interest in the rapid diagnostics industry. We report here the development of simple, low-cost optical nanosensors for the semi-quantitative detection or naked-eye detection of tyramine (a biogenic amine whose production is commonly associated with food spoilage) when coupled to Au(III)/tectomer films deposited on polylactic acid (PLA) supports. Tectomers are two-dimensional oligoglycine self-assemblies, whose terminal amino groups enable both the immobilization of Au(III) and its adhesion to PLA. Upon exposure to tyramine, a non-enzymatic redox reaction takes place in which Au(III) in the tectomer matrix is reduced by tyramine to gold nanoparticles, whose reddish-purple color depends on the tyramine concentration and can be identified by measuring the RGB coordinates (Red-Green-Blue coordinates) using a smartphone color recognition app. Moreover, a more accurate quantification of tyramine in the range from 0.048 to 10 µM could be performed by measuring the reflectance of the sensing layers and the absorbance of the characteristic 550 nm plasmon band of the gold nanoparticles. The relative standard deviation (RSD) of the method was 4.2% (n = 5) with a limit of detection (LOD) of 0.014 µM. A remarkable selectivity was achieved for tyramine detection in the presence of other biogenic amines, especially histamine. This methodology, based on the optical properties of Au(III)/tectomer hybrid coatings, is promising for its application in food quality control and smart food packaging.

Simulating Outcomes of Cataract Surgery: Important Advances in Ophthalmology.

As the human eye ages, the crystalline lens stiffens (presbyopia) and opacifies (cataract), requiring its replacement with an artificial lens [intraocular lens (IOL)]. Cataract surgery is the most frequently performed surgical procedure in the world. The increase in IOL designs has not been paralleled in practice by a sophistication in IOL selection methods, which rely on limited anatomical measurements of the eye and the surgeon's interpretation of the patient's needs and expectations. We propose that the future of IOL selection will be guided by 3D quantitative imaging of the crystalline lens to map lens opacities, anticipate IOL position, and develop fully customized eye models for ray-tracing-based IOL selection. Conversely, visual simulators (in which IOL designs are programmed in active elements) allow patients to experience prospective vision before surgery and to make more informed decisions about which IOL to choose. Quantitative imaging and optical and visual simulations of postsurgery outcomes will allow optimal treatments to be selected for a patient undergoing modern cataract surgery.

Gold nanoparticle formation as an indicator of enzymatic methods: colorimetric L-phenylalanine determination.

An enzymatic-colorimetric method has been developed based on the reaction between L-phenylalanine (L-Phe) and the L-amino acid oxidase (LAAO) in the presence of Au(III), which has led to the formation of gold nanoparticles. The intensity of the localized surface plasmon resonance (LSPR) band of the generated nanoparticles (550 nm) can be related to the concentration of L-Phe in the sample. The mechanism of the LAAO-L-Phe enzyme reaction in the presence of Au(III) has been studied through the evaluation and optimization of experimental conditions. These studies have reinforced the hypothesis that the catalytic center of the enzyme helps the Au(III) reduction and, thanks to the protein, the Au0 form is stabilized as gold nanoparticles (AuNPs). In the calibration study, a sigmoidal relationship between the concentration of the substrate and the LSPR of the nanoparticles was observed. The linearization of the signal has allowed the determination of L-Phe in the range from 17 to 500 µM with an RSD% (150 µM) of 4.8% (n = 3). The method is free of other amino acid interference normally found in blood plasma. These highly competitive results open the possibility of further development of a rapid method for L-Phe determination based on colorimetry.

Foveal and peripheral visual quality and accommodation with multifocal contact lenses.

Multifocal contact lenses are increasingly popular interventions for controlling myopia. This study presents the short-term effects of multifocal contact lenses on foveal and peripheral vision. The MiSight contact lenses designed to inhibit myopia progression and the 1-Day Acuvue Moist contact lenses designed for presbyopia were investigated. The MiSight produced similar foveal results to spectacles despite the increased astigmatism and coma. The MiSight also reduced the low-contrast resolution acuity in the periphery, despite no clear change in relative peripheral refraction. When compared with spectacles, Acuvue Moist decreased accommodative response and reduced foveal high- and low-contrast resolution acuity, whereas peripheral thresholds were more similar to those of spectacles. The most likely treatment property for myopia control by the MiSight is the contrast reduction in the peripheral visual field and the changed accommodation.

Matching convolved images to optically blurred images on the retina.

Convolved images are often used to simulate the effect of ocular aberrations on image quality, where the retinal image is simulated by convolving the stimulus with the point spread function derived from the subject's aberrations. However, some studies have shown that convolved images are perceived far more degraded than the same image blurred with optical defocus. We hypothesized that the positive interactions between the monochromatic and chromatic aberrations in the eye are lost in the convolution process. To test this hypothesis, we evaluated optical and visual quality with natural optics and with convolved images (on-bench, computer simulations, and visual acuity [VA] in subjects) using a polychromatic adaptive optics system with monochromatic (555 nm) and polychromatic light (WL) illumination. The subject's aberrations were measured using a Hartmann Shack system and were used to convolve the visual stimuli, using Fourier optics. The convolved images were seen through corrected optics. VA with convolved stimuli was lower than VA through natural aberrations, particularly in WL (by 26% in WL). Our results suggest that the systematic decrease in visual performance with visual acuity and retinal image quality by simulation with convolved stimuli appears to be primarily associated with a lack of favorable interaction between chromatic and monochromatic aberrations in the eye.

Functional integration of eye tissues and refractive eye development: Mechanisms and pathways.

Refractive eye development is a tightly coordinated developmental process. The general layout of the eye and its various components are established during embryonic development, which involves a complex cross-tissue signaling. The eye then undergoes a refinement process during the postnatal emmetropization process, which relies heavily on the integration of environmental and genetic factors and is controlled by an elaborate genetic network. This genetic network encodes a multilayered signaling cascade, which converts visual stimuli into molecular signals that guide the postnatal growth of the eye. The signaling cascade underlying refractive eye development spans across all ocular tissues and comprises multiple signaling pathways. Notably, tissue-tissue interaction plays a key role in both embryonic eye development and postnatal eye emmetropization. Recent advances in eye biometry, physiological optics and systems genetics of refractive error have significantly advanced our understanding of the biological processes involved in refractive eye development and provided a framework for the development of new treatment options for myopia. In this review, we summarize the recent data on the mechanisms and signaling pathways underlying refractive eye development and discuss new evidence suggesting a wide-spread signal integration across different tissues and ocular components involved in visually guided eye growth.

Isolated human crystalline lens three-dimensional shape: A comparison between Indian and European populations.

There have been many studies on lens properties in specific populations (e.g. in China, Europe, Singapore, etc.) some of which suggest there may be differences between populations. Differences could be caused by ethnic or environmental influences or experimental procedures. The purpose of this study is to evaluate if any differences exist between Indian and European populations in the central geometric and full shape properties of human lenses. Two custom-developed spectral domain optical coherence tomography systems were used to acquire the crystalline lens geometry: one in India (69 lenses from 59 donors) and the other in Spain (24 lenses from 19 donors). The steps for obtaining accurate 3-D models from optical coherence tomography raw images comprised of image segmentation, fan and optical distortion correction, tilt removal and registration. The outcome variables were lens equatorial diameter, lens thickness, anterior and posterior lens thicknesses and their ratio, central radius of curvature of the anterior and posterior lens surfaces, lens volume and lens surface area. A mixed effects model by maximum likelihood estimation was used to evaluate the effect of age, population and their interaction (age*population) on lens parameters. After adjusting for age, there were no population differences observed in anterior and posterior radii of curvature, equatorial diameter, lens thickness, anterior and posterior lens thicknesses and their ratio, volume and surface area (all p ≥ 0.08). There was also no effect of the interaction term on anterior and posterior radii of curvature, equatorial diameter, lens thickness, anterior and posterior lens thicknesses and their ratio, volume and surface area (all p ≥ 0.06). All central geometric and full shape parameters appeared to be comparable between the European and Indian populations. This is the first study to compare geometric and full shape lens parameters between different populations in vitro.

Smartphone-interrogated test supports for the enzymatic determination of putrescine and cadaverine in food.

Diamino-oxidase (DAO), horseradish peroxidase (HRP), and tetramethylbenzidine (TMB) have been immobilized into cellulose to obtain circular cellulose test supports (CCTSs) for the determination of cadaverine (Cad) and putrescine (Put). During the enzymatic reaction, TMB is oxidized and a blue spot is obtained. This color (RGB coordinates) is measured with a smartphone and a commercial application. The highest sensitivity is provided by the component R and a linear response is observed for low biogenic amine (BA) concentrations, but a second-order polynomial response better fits the experimental results for a wider concentration range. This has been successfully explained with a model developed to explain the RGB values obtained in this type of analytical system. Optimization studies enable CCTSs to be obtained for Put and Cad determination, which could be used (kept at 4 °C) for at least 45 days if a stabilizer (StabilCoat™ or StabilGuard™) is added during its synthesis. In these conditions, the R coordinate follows the model up to at least 4 × 10-4 M Put and/or Cad (both analytes give the same response). The method permits the Put and Cad determination from 5 × 10-5 M up to 4 × 10-4 M (RSD = 3%, n = 3). The CCTSs have been applied to Put + Cad determination in a tuna sample without any interference by other biogenic amines. The concentration found statistically agrees with that obtained using a HPLC-MS-validated method. Graphical abstract.

Crystalline lens gradient refractive index distribution in the guinea pig.

PURPOSE: The crystalline lens undergoes morphological and functional changes with age and may also play a role in eye emmetropisation. Both the geometry and the gradient index of refraction (GRIN) distribution contribute to the lens optical properties. We studied the lens GRIN in the guinea pig, a common animal model to study myopia. METHODS: Lenses were extracted from guinea pigs (Cavia porcellus) at 18 days of age (n = 4, three monolaterally treated with negative lenses and one untreated) and 39 days of age (n = 4, all untreated). Treated eyes were myopic (-2.07 D on average) and untreated eyes hyperopic (+3.3 D), as revealed using streak retinoscopy in the live and cyclopeged animals. A custom 3D spectral domain optical coherence tomography (OCT) system (λ = 840 nm, Δλ = 50 nm) was used to image the enucleated crystalline lens at two orientations. Custom algorithms were used to estimate the lens shape and GRIN was modelled with four variables that were reconstructed using the OCT data and a minimisation algorithm. Ray tracing was used to calculate the optical power and spherical aberration assuming a homogeneous refractive index or the estimated GRIN. RESULTS: Guinea pig lenses exhibited nearly parabolic GRIN profiles. When comparing the two age groups (18- and 39 day-old) there was a significant increase in the central thickness (from 3.61 to 3.74 mm), and in the refractive index of the surface (from 1.362 to 1.366) and the nucleus (from 1.443 to 1.454). The presence of GRIN shifted the spherical aberration (-4.1 µm on average) of the lens towards negative values. CONCLUSIONS: The guinea pig lens exhibits a GRIN profile with surface and nucleus refractive indices that increase slightly during the first days of life. GRIN plays a major role in the lens optical properties and should be incorporated into computational guinea pig eye models to study emmetropisation, myopia development and ageing.

VioBio lab adaptive optics: technology and applications by women vision scientists.

PURPOSE: Adaptive Optics allows measurement and manipulation of the optical aberrations of the eye. We review two Adaptive Optics set-ups implemented at the Visual Optics and Biophotonics Laboratory, and present examples of their use in better understanding of the role of optical aberrations on visual perception, in normal and treated eyes. RECENT FINDINGS: Two systems (AOI and AOII) are described that measure ocular aberrations with a Hartmann-Shack wavefront sensor, which operates in closed-loop with an electromagnetic deformable mirror, and visual stimuli are projected in a visual display for psychophysical measurements. AOI operates in infrared radiation (IR) light. AOII is provided with a supercontiniuum laser source (IR and visible wavelengths), additional elements for simulation (spatial light modulator, temporal multiplexing with optotunable lenses, phase plates, cuvette for intraocular lenses-IOLs), and a double-pass retinal camera. We review several studies undertaken with these AO systems, including the evaluation of the visual benefits of AO correction, vision with simulated multifocal IOLs (MIOLs), optical aberrations in pseudophakic eyes, chromatic aberrations and their visual impact, and neural adaptation to ocular aberrations. SUMMARY: Monochromatic and chromatic aberrations have been measured in normal and treated eyes. AO systems have allowed understanding the visual benefit of correcting aberrations in normal eyes and the adaptation of the visual system to the eye's native aberrations. Ocular corrections such as intraocular and contact lenses modify the wave aberrations. AO systems allow simulating vision with these corrections before they are implanted/fitted in the eye, or even before they are manufactured, revealing great potential for industry and the clinical practice. This review paper is part of a special issue of Ophthalmic & Physiological Optics on women in visual optics, and is co-authored by all women scientists of the research team.

Colorimetric-enzymatic determination of tyramine by generation of gold nanoparticles.

In this paper, it has been demonstrated that Au(III) is able to act instead of O2 in the oxidase enzymatic reaction, so that it becomes reduced to purple gold nanoparticles (AuNPs). The plasmon band (at 540 nm) can be used as the analytical signal. Tyramine has been determined using its enzymatic reaction with tyramine oxidase (TAO). The kinetic of the AuNP formation has been studied in the light of both the Avrami equation for crystallization and the Finke-Watsy mechanism for AuNP nucleation and growth. The effects of the Au(III), TAO and tyramine concentrations on the corresponding kinetic constants have been investigated. Working at room temperature, under optimal conditions (phosphate buffer pH 7.0, TAO 0.5 U.mL-1 Au(III) 1 mM), the linear response ranges from 2.5 × 10-5 M to 3.3 × 10-4 M Tyramine (5.6% RSD) and the LOD is 2.9 × 10-6 M. Under these conditions, the signal is measured after 30 min reaction (to obtain the highest sensitivity), but this time can be significantly reduced by increasing the temperature (the reaction is finished after 4 min when working at 50 °C). The method has been applied to tyramine determination in a cheese sample with good results. The new scheme proposed in this paper can be extended, in principle, to other enzymatic methods based on oxidase enzymes. Graphical abstractTyramine is determined by measuring the plasmon band of the gold nanoparticles formed during its enzymatic reaction with Tyramine oxidase. Moreover, a mathematical model has been developed to explain the formation of the gold nanoparticles during the reaction.

Tunable lenses: dynamic characterization and fine-tuned control for high-speed applications.

Tunable lenses are becoming ubiquitous, in applications including microscopy, optical coherence tomography, computer vision, quality control, and presbyopic corrections. Many applications require an accurate control of the optical power of the lens in response to a time-dependent input waveform. We present a fast focimeter (3.8 KHz) to characterize the dynamic response of tunable lenses, which was demonstrated on different lens models. We found that the temporal response is repetitive and linear, which allowed the development of a robust compensation strategy based on the optimization of the input wave, using a linear time-invariant model. To our knowledge, this work presents the first procedure for a direct characterization of the transient response of tunable lenses and for compensation of their temporal distortions, and broadens the potential of tunable lenses also in high-speed applications.

Gold nanoclusters as a quenchable fluorescent probe for sensing oxygen at high temperatures.

Gold nanoclusters (AuNCs) capped with lipoic acid (LA) or templated with bovine serum albumin (BSA) are shown to be viable fluorescent probes for oxygen (O2) which acts as a collisional quencher. Quenching of fluorescence, with its lifetimes in the order of 123 ± 9 ns (LA) and 153 ± 15 ns (BSA) (in aqueous solution), is best measured at excitation/emission wavelengths of 400/680 nm and 375/650 nm respectively. It follows the Stern-Volmer model, whose quenching constants (Ksv) and quenching efficiencies (γ) are 1400 M-1 and 0.52 for AuNC@LA and 4479 M-1 and 0.90 for AuNC@BSA. The probes were immobilized on a silica support and tested for response to O2 in gas phase using a commercial instrument. The effect of temperature on the fluorescence of AuNC@LA was studied in the range from 30 to 210 °C. Fluorescence intensity slightly decreases with temperature in the first heating cycle but remains constant in further cycles. The AuNC@LA were studied for their response to O2 in the temperature range from 30 to 100 °C, and even at 100 °C they respond to O2, with a Ksv that slightly drops with increasing temperature. Measuring in gas phase at 100 °C, the sensor has a detection limit of 3% (V/V) of O2 at a signal-to-noise ratio of 3. Graphical Abstract Gold-nanoclusters (AuNCs) fluorescence intensity (λexc = 400 nm, λem = 680 nm) remains constant from 30 to 210 °C and is quenched by O2 following a collisional mechanism. The Stern-Volmer constant (Ksv) slightly changes from 25 °C to 100 °C (at least).

Virtual pyramid wavefront sensor for phase unwrapping.

Noise affects wavefront reconstruction from wrapped phase data. A novel method of phase unwrapping is proposed with the help of a virtual pyramid wavefront sensor. The method was tested on noisy wrapped phase images obtained experimentally with a digital phase-shifting point diffraction interferometer. The virtuality of the pyramid wavefront sensor allows easy tuning of the pyramid apex angle and modulation amplitude. It is shown that an optimal modulation amplitude obtained by monitoring the Strehl ratio helps in achieving better accuracy. Through simulation studies and iterative estimation, it is shown that the virtual pyramid wavefront sensor is robust to random noise.

From unseen to seen: tackling the global burden of uncorrected refractive errors.

Worldwide, more than one billion people suffer from poor vision because they do not have the eyeglasses they need. Their uncorrected refractive errors are a major cause of global disability and drastically reduce productivity, educational opportunities, and overall quality of life. The problem persists most prevalently in low-resource settings, even though prescription eyeglasses serve as a simple, effective, and largely affordable solution. In this review, we discuss barriers to obtaining, and approaches for providing, refractive eye care. We also highlight emerging technologies that are being developed to increase the accessibility of eye care. Finally, we describe opportunities that exist for engineers to develop new solutions to positively impact the diagnosis and treatment of correctable refractive errors in low-resource settings.