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In hereditary spherocytosis (HS), genetic mutations in the cell membrane and cytoskeleton proteins cause structural defects in red blood cells (RBCs). As a result, cells are rigid and misshapen, usually with a characteristic spherical form (spherocytes), too stiff to circulate through microcirculation regions, so they are prone to undergo hemolysis and phagocytosis by splenic macrophages. Mild to severe anemia arises in HS, and other derived symptoms like splenomegaly, jaundice, and cholelithiasis. Although abnormally shaped RBCs can be identified under conventional light microscopy, HS diagnosis relies on several clinical factors and sometimes on the results of complex molecular testing. It is specially challenging when other causes of anemia coexist or after recent blood transfusions. We propose two different approaches to characterize RBCs in HS: (i) an immunofluorescence assay targeting protein band 3, which is affected in most HS cases and (ii) a three-dimensional morphology assay, with living cells, staining the membrane with fluorescent dyes. Confocal laser scanning microscopy (CLSM) was used to carry out both assays, and in order to complement the latter, a software was developed for the automated detection of spherocytes in blood samples. CLSM allowed the precise and unambiguous assessment of cell shape and protein expression. This publication is part of the project PID2020-112527RBI00, funded by CIN/AEI/10.13039/501100011033. Laura Rey-Barroso thanks the Ministry of Science, Innovation and Universities for the PhD (FPI) grant she has received (DPI2017-89414-R). The current study has been funded by the Spanish National Agency of Investigation (AEI).

Hemoglobinopathies represent the most common single-gene defects in the world and pose a major public health problem. These defects can affect the synthesis of hemoglobin, the molecule in charge of oxygen transportation, quantitatively (thalassemia) or qualitatively (sickle cell disease). These affectations will terminally lead to the premature destruction of red blood cells (RBCs), nevertheless they have different etiologies and they need to be treated differently. Diagnosing a patient can sometimes be difficult due to the coexistence of several of such diseases and/or blood transfusions, and requires expensive and complex molecular tests. But alternatively, it is well-known that the spectral information of RBC autofluorescence and immunostaining can reveal important differences among these RBCs diseases [1] – [3] . The isolated assessment of cell shape has been proven to be insufficient. In these context, we conducted a first pilot study to analyze the spectral and morphological characteristics of healthy and diseased RBCs, under a spectral confocal microscope Leica TCS SP8 (Leica Microsystems GmbH, Mannheim, Germany). Specifically, fresh blood samples from 17 pediatric patients ages 1 to 17, some suffering from α-thalassemia and iron deficiency were evaluated. Autofluorescence was captured in the 425-790 nm range, using hybrid detection that provides with a higher sensitivity, a 63x (NA 1.4, oil) plan-apochromatic objective, and a diode laser line with emission at 405 nm for excitation. The autofluorescence signals captured showed spectral differences that can be used to discriminate among diseases and severity, especially at the 628 nm band as showed in Figure 1 .

Purpose: To develop a methodology based on a double-pass system to obtain information about the transmittance of ocular media. Methods: The procedure consists of recording double-pass images at different powers of a laser diode of 780 nm and determining the scattering in an area between 25–35 arcmin of each image. The scattering showed linear behavior in respect to the irradiance of the laser, and the slope of the linear fit was proportional to the transmittance squared of the media evaluated. An artificial eye with different filters was tested first. Then, fifteen subjects with clear ocular media were divided into two groups: ten subjects classified by the iris color were recruited for the measurements of an ocular transmittance index and the estimation of the transmittance (group A), and another five subjects were selected for measurements with neutral filters (group B). Results: The measurements performed in group A presented a mean transmittance of 42.95%. No differences in the transmittance were found between subjects with different iris color (p = 0.154). Measurements in group B showed a good correlation (r = 0.959, p < 0.001) between the expected and the measured value for the transmittance. Conclusion: We proposed and evaluated a method to determine the transmittance of the eye in vivo using the double-pass system.

Decreasing the use of pesticides is one of the main goals of current agriculture, which requires fast, precise and continuous assessments of crop pests. Citrus pests cause a lot of damage worldwide and the techniques to evaluate them are mainly based on manual, time-consuming readings of insects stuck on traps spread over the crops. This is the case of red scale insects, whose control is notably challenging due to their small size and high reproduction rate. Hence, in this work, we carry out a spectral characterization of this insect in the visible range through spectrometric devices, microscopy and hyperspectral imaging technology to analyze the feasibility of using this information as a means of automatically identifying specimens belonging to this species in this era of precision agriculture. The results obtained show that spectral reflectance differences between red scales and other insects can be recorded at long (red) wavelengths and that red scales are morphologically different, i.e., smaller and more rounded. A reflectance ratio computed from spectral images taken at 774 nm and 410 nm is proposed as a new approach for automated discrimination of red scales from other insects. This project has been co-financed by the European Union through the European Regional Development Fund (ERDF) and has the support of the Secretariat of Universities and Research of the Department of Business and Knowledge of the Generalitat de Catalunya (Exp. 2019PROD00013)

Sparkle and graininess are textural effects that appear as intrinsic spatial features of coatings containing goniochromatic pigments, whereas others such as mottling are undesired outcomes. In this study, we present new methods for the evaluation of sparkle, graininess and mottling of automotive coatings through images acquired with a novel gonio-hyperspectral imaging system based on light-emitting diodes with extended spectral sensitivity beyond the visible range (368&ndash, 1309 nm). A novel analysis of sparkle that considered the amount of sparkling spots was used. Graininess was quantified through a novel statistical descriptor based on the energy descriptor computed from the histogram. A new index was proposed for an enhanced detection of mottling based on the analysis of striping, which was better observed and quantified in the infrared range. Spectral assessments showed a strong relationship with color and spectral reflectance for graininess and mottling. In conclusion, the results showed that these new spatial and spectral methods are a suitable, alternative and potential approach for the assessment of these textural features.

We present a multispectral fundus camera that performs fast imaging of the ocular posterior pole in the visible and near-infrared (400 to 1300 nm) wavelengths through 15 spectral bands, using a flashlight source made of light-emitting diodes, and CMOS and InGaAs cameras. We investigate the potential of this system for visualizing occult and overlapping structures of the retina in the unexplored wavelength range beyond 900 nm, in which radiation can penetrate deeper into the tissue. Reflectance values at each pixel are also retrieved from the acquired images in the analyzed spectral range. The available spectroscopic information and the visualization of retinal structures, specifically the choroidal vasculature and drusen-induced retinal pigment epithelium degeneration, which are hardly visible in conventional color fundus images, underline the clinical potential of this system as a new tool for ophthalmic diagnosis.

The use of terahertz-based techniques has grown very fast since they are capable of performingevaluations at molecular level, being very suitable for the analysis of biological samples and bioma-terials such as those for contact lenses. These biomaterials are continuously evolving to enhancethe lens wearer’s comfort by improving their hydration state and surface wettability. Therefore, thisstudy examines a novel terahertz system for the assessment of the temporary in vitro dehydrationof hydrogel contact lenses, which provides a new index to assess their state of hydration. Severalconventional and silicone hydrogel contact lenses and lens care solutions were analysed. Traditionalmethods such as the gravimetric determination of water content and the measurement of the staticcontact angle were also carried out for the validation of the developed system. The dehydrationrate measurements of contact lenses obtained with the proposed system correlated with the valuesprovided by traditional methods. As a whole, conventional hydrogel contact lenses exhibited thelowest values for dehydration rate. The tests conducted on various solutions showed a correlationbetween the wetting action of the solution and the dehydration rate of the contact lens material.

The effective and non-invasive diagnosis of skin cancer is a hot topic, since biopsy is a costly and time-consuming surgical procedure. As skin relief is an important biophysical feature that can be difficult to perceive with the naked eye and by touch, we developed a novel 3D imaging scanner based on fringe projection to obtain morphological parameters of skin lesions related to perimeter, area and volume with micrometric precision. We measured 608 samples and significant morphological differences were found between melanomas and nevi (p

The process of separating small vertebrate fossils from sediment is a monotonous exercise, usually carried out manually and mainly based on morphological features. The large amount of time and personnel required results in the high economic costs associated with this antiquated technique. In this study we test the potential of colour and multispectral imaging for the detection of small vertebrate fossils to facilitate their separation from sediment. This technique is based on the analysis of the spectral and fluorescence characteristics of fossils. Specifically, a multispectral system with a charge-coupled device (CCD) camera attached to a liquid crystal tunable filter and a digital colour camera was used in combination with daylight and ultraviolet light sources. The results show that the reflectance of bones, teeth and sediment are usually different at longer wavelengths, and that fluorescence rates also differ in the bluegreen region. Unfortunately, samples from different paleontological sites present different patterns, hindering the standardization of a method that discriminates microfossils from sediments. However, the combination of colour and multispectral imaging systems with recent advances in laser-stimulated fluorescence in fossils could constitute an optimal solution for an efficient sorting process of small vertebrate remains within geological samples.

In this study, a novel gonio-hyperspectral imaging system based on light-emitting diodes for the analysis of automotive coatings was validated colorimetrically and spectrally from 368 to 1309 nm. A total of 30 pearlescent, 30 metallic, and 30 solid real automotive coatings were evaluated with this system, the BYK-mac and X-Rite MA98 gonio-spectrophotometers, and also with the SPECTRO 320 spectrometer for further comparison. The results showed very precise correlations, especially in the visible range. In conclusion, this new system provides a deeper assessment of goniochromatic pigments than current approaches due to the expansion of the spectral range to the infrared. Ministerio de Ciencia e Innovación (MICINN) (DPI2011-30090-C02-01); European Union. Francisco J. Burgos-Fernández thanks the Government of Catalonia for his Ph.D. grant.

The effective and non-invasive diagnosis of skin cancer is a hot topic in biophotonics since the current gold standard, biopsy followed by histological examination, is a slow and costly procedure for the healthcare system. Therefore, authors have put their efforts in characterizing skin cancer quantitatively through optical and photonic techniques such as 3D topography and multispectral imaging. Skin relief is an important biophysical feature that can be difficult to appreciate by touch, but can be precisely characterized with 3D imaging techniques, such as fringe projection. Color and spectral features given by skin chromophores, which are routinely analyzed by the naked eye and through dermoscopy, can also be quantified by means of multispectral imaging systems. In this study, the outcomes of these two imaging modalities were combined in a machine learning process to enhance classification of melanomas and nevi obtained from the two systems when operating isolately. The results suggest that the combination of 3D and multispectral data is relevant for the medical diagnosis of skin cancer.

With the goal of diagnosing skin cancer in an early and noninvasive way, an extended near infrared multispectral imaging system based on an InGaAs sensor with sensitivity from 995 nm to 1613 nm was built to evaluate deeper skin layers thanks to the higher penetration of photons at these wavelengths. The outcomes of this device were combined with those of a previously developed multispectral system that works in the visible and near infrared range (414 nm–995 nm). Both provide spectral and spatial information from skin lesions. A classification method to discriminate between melanomas and nevi was developed based on the analysis of first-order statistics descriptors, principal component analysis, and support vector machine tools. The system provided a sensitivity of 78.6% and a specificity of 84.6%, the latter one being improved with respect to that offered by silicon sensors.

The effective and non-invasive diagnosis of skin cancer is a hot topic in biophotonics since the current gold standard, a biopsy, is slow and costly. Non-invasive optical techniques such as polarization and multispectral imaging have arisen as powerful tools to overcome these constraints. The combination of these techniques provides a comprehensive characterization of skin chromophores including polarization, color and spectral features. Hence, in this work we propose a polarized multispectral imaging device that works from 414 nm to 995 nm and at 0º, 45º and 90º polarization configurations. Preliminary results performed over 20 nevi and 20 melanoma found statistically significant descriptors (p