A Performance Comparison of 3D Survey Instruments for Their Application in the Cultural Heritage Field.

Thanks to the recent development of innovative instruments and software with high accuracy and resolution, 3D modelling provides useful insights in several sectors (from industrial metrology to cultural heritage). Moreover, the 3D reconstruction of objects of artistic interest is becoming mandatory, not only because of the risks to which works of art are increasingly exposed (e.g., wars and climatic disasters) but also because of the leading role that the virtual fruition of art is taking. In this work, we compared the performance of four 3D instruments based on different working principles and techniques (laser micro-profilometry, structured-light topography and the phase-shifting method) by measuring four samples of different sizes, dimensions and surface characteristics. We aimed to assess the capabilities and limitations of these instruments to verify their accuracy and the technical specifications given in the suppliers' data sheets. To this end, we calculated the point densities and extracted several profiles from the models to evaluate both their lateral (XY) and axial (Z) resolution. A comparison between the nominal resolution values and those calculated on samples representative of cultural artefacts was used to predict the performance of the instruments in real case studies. Overall, the purpose of this comparison is to provide a quantitative assessment of the performance of the instruments that allows for their correct application to works of art according to their specific characteristics.

The Degradation of Daguerreotypes and the Relationship with Their Multi-Material Structure: A Multimodal Investigation.

Preserving and analytically examining daguerreotypes is particularly challenging because of their multi-material and multi-component structure. Various sensors have been exploited to examine mainly the image plates of the daguerreotypes even though the degradation goes beyond this component. Micro-analyses have been the preferred method due to the nanoscale structure of the image particles. In this work, we propose comprehensive multi-modal non-invasive sensing to investigate the corrosion products present in nine daguerreotypes from the Fondazione Alinari per la Fotografia (FAF, Florence, Italy). The methodology proposed includes chemical and morphological analyses: portable X-ray fluorescence spectrometry (pXRF), Raman microspectroscopy (µ-Raman), and micro-Fourier transform infrared spectroscopy in reflection mode (µ-rFTIR) for the chemical identification. For the first time, optical coherence tomography (OCT) was deployed to record the cross-sectional and morphological data of the relevant corrosion formations on daguerreotypes in a contactless way. The results allowed the characterization, in a non-invasive mode at a microscopic level, of a wide range of degradation products produced by the interaction of the different elements present in the structure of the daguerreotypes. The aim was to verify the performance of the proposed methodology and to link the chemical and physical complexity of the entire structure, disclosed by the state-of-art sensors, to the daguerreotype degradation. The results draw attention to the need to monitor not only the image condition but the whole object as a partially closed system in constant interaction internally and with the environment.

Reflectance Spectroscopy as a Novel Tool for Thickness Measurements of Paint Layers.

A major challenge in heritage science is the non-invasive cross-sectional analysis of paintings. When low-energy probes are used, the presence of opaque media can significantly hinder the penetration of incident radiation, as well as the collection of the backscattered signal. Currently, no technique is capable of uniquely and noninvasively measuring the micrometric thickness of heterogeneous materials, such as pictorial layers, for any painting material. The aim of this work was to explore the possibility of extracting stratigraphic information from reflectance spectra obtained by diffuse reflectance spectroscopy (DRS). We tested the proposed approach on single layers of ten pure acrylic paints. The chemical composition of each paint was first characterised by micro-Raman and laser-induced breakdown spectroscopies. The spectral behaviour was analysed by both Fibre Optics Reflectance Spectroscopy (FORS) and Vis-NIR multispectral reflectance imaging. We showed that there is a clear correlation between the spectral response of acrylic paint layers and their micrometric thickness, which was previously measured by Optical Coherence Tomography (OCT). Based on significant spectral features, exponential functions of reflectance vs. thickness were obtained for each paint, which can be used as calibration curves for thickness measurements. To the best of our knowledge, similar approaches for cross-sectional measurements of paint layers have never been tested.

Portable Sequentially Shifted Excitation Raman Spectroscopy to Examine Historic Powders Enclosed in Glass Vials.

Raman spectroscopy (RS) is a powerful non-invasive tool for the characterization of materials. However, the fluorescence effect often hampers the detectability of the relatively weak vibrational Raman signal. Several approaches were exploited to overcome this limit. This work, in particular, evaluates the performance of an in situ portable sequentially shifted excitation (SSE™) Raman spectrometer applied to the examination of artistic historical pigment powders enclosed in glass vials. The explored handheld spectrometer employs a dual, temperature-shifted, 785 nm and 852 nm laser excitation to optimize both spectral coverage and fluorescence subtraction. The study demonstrates the feasibility of the SSE RS approach for non-invasive identification of art materials, and its applicability in complex situations where the examined material cannot be removed from its container. Laboratory measurements using benchtop dispersive micro-Raman spectroscopy at 785 nm are reported for comparison.

Correction: Dal Fovo et al. Multi-Analytical Characterization and Radiocarbon Dating of a Roman Egyptian Mummy Portrait. Molecules 2021, 26, 5268.

Due to the fact that the policy regarding the publication of images from the collection of the Papyrological Institute, the owner of the object under study, changed when the article was already in publication, the authors would like to make the following corrections to this paper [...].

Automated Phasor Segmentation of Fluorescence Lifetime Imaging Data for Discriminating Pigments and Binders Used in Artworks.

The non-invasive analysis of fluorescence from binders and pigments employed in mixtures in artworks is a major challenge in cultural heritage science due to the broad overlapping emission of different fluorescent species causing difficulties in the data interpretation. To improve the specificity of fluorescence measurements, we went beyond steady-state fluorescence measurements by resolving the fluorescence decay dynamics of the emitting species through time-resolved fluorescence imaging (TRFI). In particular, we acquired the fluorescence decay features of different pigments and binders using a portable and compact fibre-based imaging setup. Fluorescence time-resolved data were analysed using the phasor method followed by a Gaussian mixture model (GMM) to automatically identify the populations of fluorescent species within the fluorescence decay maps. Our results demonstrate that this approach allows distinguishing different binders when mixed with the same pigment as well as discriminating different pigments dispersed in a common binder. The results obtained could establish a framework for the analysis of a broader range of pigments and binders to be then extended to several other materials used in art production. The obtained results, together with the compactness and portability of the instrument, pave the way for future in situ applications of the technology on paintings.

Synchronized Hardware-Registered VIS-NIR Imaging Spectroscopy and 3D Sensing on a Fresco by Botticelli.

We discuss a synchronised sensing technique for the analysis of painted surfaces of frescos. Specifically, the performance of Visible-Near Infrared (VIS-NIR) Reflectance Imaging Spectroscopy (RIS) synchronized with three-dimensional (3D) acquisition is demonstrated in the study of a detached mural painting by Alessandro Botticelli. Synchronized sensing generates georeferenced data for simplified data treatment and interpretation. We show how such output data can provide key information to interpret important fresco surface and subsurface features (e.g., painting technique, material composition, pentimenti).

Multisensorial Assessment of Laser Effects on Shellac Applied on Wall Paintings.

The assessment of five different laser treatments in the conservation of wall paintings was devised on the basis of the surface temperature monitoring by infrared thermography (IRT), ultraviolet-induced fluorescence-visible (UV-VIS) imaging, and optical coherence tomography (OCT). A series of yttrium-aluminum-garnet (YAG) lasers were tested for removal of shellac layers from wall painting mock-ups. The mock-ups were realized as buon fresco with different mineral based pigments (earths and iron oxide) on a lime- and sand-based mortar. After the carbonatation process, all the samples were treated with shellac (5% in ethanol). The effects of neodymium (Nd):YAG, holmium (Ho):YAG, and erbium (Er):YAG laser sources, in different operative modes, on average temperature of the surface, color, and morphology were inspected with complementary sensors. The results show the necessity to adopt a combined approach in establishing safe laser operating conditions to avoid any undesired effects induced on the artefacts by the laser treatments. We demonstrate, for the first time, the performance of the Ho:YAG laser in the removal of a conservation treatment.

Multi-Analytical Characterization and Radiocarbon Dating of a Roman Egyptian Mummy Portrait.

Fayum mummy portraits, painted around 2000 years ago, represent a fascinating fusion of Egyptian and Graeco-Roman funerary and artistic traditions. Examination of these artworks may provide insight into the Roman Empire's trade and economic and social structure during one of its most crucial yet still hazy times of transition. The lack of proper archaeological documentation of the numerous excavated portraits currently prevents their chronological dating, be it absolute or relative. So far, their production period has been defined essentially on the basis of the relevant differences in their pictorial style. Our study introduces the use of Accelerator Mass Spectrometry (AMS) to assess the age of a fragment of an encaustic painting belonging to the corpus of the Fayum portraits. The unexpected age resulting from 14C analysis suggests the need to reconsider previous assumptions regarding the period of production of the Fayum corpus. Furthermore, our multi-analytical, non-invasive approach yields further details regarding the fragment's pictorial technique and constituting materials, based on spectral and morphological analysis and cross-sectional examination.

Combined photoacoustic imaging to delineate the internal structure of paintings.

In this Letter, we present a combined photoacoustic imaging method, based on consecutive excitation using either the fundamental or the second-harmonic wavelength of a pulsed Nd:YAG laser for the stratigraphy of painted artworks. Near-infrared excitation was employed for the imaging of hidden underdrawings in mock-up samples, whereas visible light was used for the thickness mapping of the overlying paint through the detection of photoacoustic signal attenuation. The proposed methodology was proven effective in measuring thick and strongly absorbing layers, which would not be possible by means of other pure optical techniques, while also enabling the visualization of features underneath the painted surface. Such an implementation expands significantly the applicability of the previously presented photoacoustic technique, which was limited to point-measurements, and paves the way for novel application in historical and technical studies, as well as in documenting restoring operations.

Using Optical Coherence Tomography to Reveal the Hidden History of The Landsdowne Virgin of the Yarnwinder by Leonardo da Vinci and Studio.

Optical coherence tomography (OCT) was used for non-invasive examination of a well-known, yet complex, painting from the studio of Leonardo da Vinci in combination with routine imaging in various bands of electromagnetic radiation. In contrast with these techniques, OCT provides depth-resolved information. Three post-processing modalities were explored: cross-sectional views, maps of scattering from given depths, and their 3D models. Some hidden alterations of the painting owing to past restorations were traced: retouching and overpainting with their positioning within varnish layers as well as indications of a former transfer to canvas.

Spectral Imaging and Archival Data in Analysing Madonna of the Rabbit Paintings by Manet and Titian.

A concise insight into the outputs provided by the latest prototype of visible-near infrared (Vis-NIR) multispectral scanner (National Research Council-National Institute of Optics, CNR-INO, Italy) is presented. The analytical data acquired on an oil painting Madonna of the Rabbit by É. Manet are described. In this work, the Vis-NIR was complemented with X-ray fluorescence (XRF) mapping for the chemical and spatial characterization of several pigments. The spatially registered Vis-NIR data facilitated their processing by spectral correlation mapping (SCM) and artificial neural network (ANN) algorithm, respectively, for pigment mapping and improved visibility of pentimenti and of underdrawing style. The data provided several key elements for the comparison with a homonymous original work by Titian studied within the ARCHive LABoratory (ARCHLAB) transnational access project.

Close to the diffraction limit in high resolution ATR FTIR mapping: demonstration on micrometric multi-layered art systems.

This paper is aimed at demonstrating the potentiality of high resolution Attenuated Total Reflection Fourier Transform Infrared micro-mapping (micro-ATR-FTIR) to reconstruct the images of micrometric multi-layered systems. This method can be an effective analytical alternative when the layer thickness requires high lateral resolution, and fluorescence or thermal effects prevent the deployment of conventional analytical techniques such as micro-Raman spectroscopy. This study demonstrates the high micro-ATR-FTIR setup performances in terms of lateral resolution, spectral quality and chemical image contrast using a new laboratory instrument equipped with a single element detector. The method has been first validated on mock-ups and then successfully applied on cross-sectional samples from real artworks: Leonardo da Vinci's mural painting, characterised by a few micrometers thin sequence of organic and inorganic layers, and an outdoor marble statue, with a complex sequence of decay products on its surface. This study paves the way to a new investigation modality of micrometric systems, combining high lateral resolution with excellent spectral quality, essential in the field of Cultural Heritage as well as in the wider area of materials and forensic sciences.

Cholesterol metabolites and tumor microenvironment: the road towards clinical translation.

Targeting the tumor microenvironment focusing on immune cells has recently become a standard of care for some tumors. Indeed, antibodies blocking immune checkpoints (e.g., anti-CTLA-4 and anti-PD1 mAbs) have been approved by regulatory agencies for the treatment of some solid tumors based upon successes in many clinical trials. Although tumor metabolism has always attracted the attention of tumor biologists, only recently have oncologists renewed their interest in this field of tumor biology research. This has highlighted the possibility to pharmacologically target rate-limiting enzymes along key metabolic pathways of tumor cells, such as lipogenesis and aerobic glycolysis. Altered tumor metabolism has also been shown to influence the functionality of the tumor microenvironment as a whole, particularly the immune cell component of thereof. Cholesterol, oxysterols and Liver X receptors (LXRs) have been investigated in different tumor models. Recent in vitro and in vivo results point to their involvement in tumor and immune cell biology, thus making the LXR/oxysterol axis a possible target for novel antitumor strategies. Indeed, the possibility to target both tumor cell metabolism (i.e., cholesterol metabolism) and tumor-infiltrating immune cell dysfunctions induced by oxysterols might result in a synergistic antitumor effect generating long-lasting memory responses. This review will focus on the role of cholesterol metabolism with particular emphasis on the role of the LXR/oxysterol axis in the tumor microenvironment, discussing mechanisms of action, pros and cons, and strategies to develop antitumor therapies based on the modulation of this axis.

The administration of drugs inhibiting cholesterol/oxysterol synthesis is safe and increases the efficacy of immunotherapeutic regimens in tumor-bearing mice.

Tumor-derived metabolites dampen tumor-infiltrating immune cells and antitumor immune responses. Among the various metabolites produced by tumors, we recently showed that cholesterol oxidized products, namely oxysterols, favor tumor growth through the inhibition of DC migration toward lymphoid organs and by promoting the recruitment of pro-tumor neutrophils within the tumor microenvironment. Here, we tested different drugs capable of blocking cholesterol/oxysterol formation. In particular, we tested efficacy and safety of different administration schedules, and of immunotherapy-based combination of a class of compounds, namely zaragozic acids, which inhibit cholesterol pathway downstream of mevalonate formation, thus leaving intact the formation of the isoprenoids, which are required for the maturation of proteins involved in the immune cell function. We show that zaragozic acids inhibit the in vivo growth of the RMA lymphoma and the Lewis lung carcinoma (LLC) without inducing side effects. Tumor growth inhibition requires an intact immune system, as immunodeficient tumor-bearing mice do not respond to zaragozic acid treatment. Of note, the effect of zaragozic acids is accompanied by a marked reduction in the LXR target genes Abcg1, Mertk, Scd1 and Srebp-1c in the tumor microenvironment. On the other hand, zoledronate, which blocks also isoprenoid formation, did not control the LLC tumor growth. Finally, we show that zaragozic acids potentiate the antitumor effects of active and adoptive immunotherapy, significantly prolonging the overall survival of tumor-bearing mice treated with the combo zaragozic acids and TAA-loaded DCs. This study identifies zaragozic acids as new antitumor compounds exploitable for the treatment of cancer patients.

Inhibition of CCR7/CCL19 axis in lesional skin is a critical event for clinical remission induced by TNF blockade in patients with psoriasis.

Despite the evidence that tumor necrosis factor (TNF) inhibitors block TNF and the downstream inflammatory cascade, their primary mechanism of action in inhibiting the self-sustaining pathogenic cycle in psoriasis is not completely understood. This study has the aim to identify early critical events for the resolution of inflammation in skin lesions using anti-TNF therapy. We used a translational approach that correlates gene expression fold change in lesional skin with the Psoriasis Area and Severity Index score decrease induced by TNF blockade after 4 weeks of treatment. Data were validated by immunofluorescence microscopy on skin biopsy specimens. We found that the anti-TNF-modulated genes that mostly associated with the clinical amelioration were Ccr7, its ligand, Ccl19, and dendritic cell maturation genes. Decreased expression of T-cell activation genes and Vegf also associated with the clinical response. More important, the down-regulation of Ccr7 observed at 4 weeks significantly correlated with the clinical remission occurring at later time points. Immunofluorescence microscopy on skin biopsy specimens showed that reduction of CCR7(+) cells and chemokine ligand (CCL) 19 was paralleled by disaggregation of the dermal lymphoid-like tissue. These data show that an early critical event for the clinical remission of psoriasis in response to TNF inhibitors is the inhibition of the CCR7/CCL19 axis and support its role in psoriasis pathogenesis.

Clinical and immunologic responses in melanoma patients vaccinated with MAGE-A3-genetically modified lymphocytes.

Cancer vaccines have recently been shown to induce some clinical benefits. The relationship between clinical activity and anti-vaccine T cell responses is somewhat controversial. Indeed, in many trials it has been documented that the induction of vaccine-specific T cells exceeds the clinical responses observed. Here, we evaluate immunological and clinical responses in 23 MAGE-A3(+) melanoma patients treated with autologous lymphocytes genetically engineered to express the tumor antigen MAGE-A3 and the viral gene product thymidine kinase of the herpes simplex virus (HSV-TK). HSV-TK was used as safety system in case of adverse events and as tracer antigen to monitor the immune competence of treated patients. The increase of anti-TK and anti-MAGE-A3 T-cells after vaccination was observed in 90 and 27% of patients, respectively. Among 19 patients with measurable disease, we observed a disease control rate of 26.3%, with one objective clinical response, and four durable, stable diseases. Three patients out of five with no evidence of disease (NED) at the time of vaccination remained NED after 73+, 70+ and 50+ months. Notably, we report that only patients experiencing MAGE-A3-specific immune responses showed a clinical benefit. Additionally, we report that responder and non-responder patients activate and expand T cells against the tracer antigen TK in a similar way, suggesting that local rather than systemic immune suppression might be involved in limiting clinically relevant antitumor immune responses.

Nonlinear imaging and vibrational spectroscopic analysis of cellulosic fibres treated with COEX® flame-retardant for tapestry preservation.

Cellulose-based fabrics are widely used in the preservation and storage of historic tapestries. Their ease of flammability is a serious concern that greatly limits their applications and requires the development of effective and safe flame-retardant treatments. In this work, we analysed linen and cotton textile samples before and after COEX® treatment, a patented green technology imparting anti-flame properties by functionalizing the cellulose molecules with phosphorus and sulphur groups. Some of the samples were also exposed to photo-induced ageing after the treatment. The resulting structural and chemical changes in both fibres were characterized by nonlinear optical imaging modalities, namely Second Harmonic Generation (SHG) and Two-Photon Excited Fluorescence microscopies (TPEF), and Raman and Attenuated Total Reflection - Fourier Transform Infrared (ATR-FTIR) spectroscopies. Complementary results evidenced a reduction in microfibril crystallinity, attributed mainly to the reduction of hydrogen bonding among cellulose macromolecules, with a concomitant increase in fluorescence possibly due to the introduction of ester groups into cellulose chains and to decomposition of lignin into fluorescent by-products.

Non-invasive contactless analysis of an early drawing by Raffaello Sanzio by means of optical methods.

Studying highly valuable and fragile Renaissance drawings requires a non-invasive contactless analytical approach. In this work, we study an early drawing by Raffaello Sanzio, one of the most important artists of the Italian Renaissance, realized during his stay in Florence (1504-1508). Our analyses aimed to reveal the features of the paper support significant for its dating, identify and map the drawing media to understand the artist's technical approach, and document the drawing condition with imaging and single-sited optical methods. Reflectance Vis-NIR multispectral imaging spectroscopy elaborated with False-Colour and Principal Component Analysis provided information about the paper support and the drawing media. Laser scanning micro-profilometry and Optical Coherence Tomography allowed revealing the superficial micro-scale features of the support. The chemical composition of the different drawing materials was characterized by µ-Raman spectroscopy, which provided also some evidence of the conservation history of the drawing. Integration of spectroscopic and imaging data shows that Raffaello used different dry drawing media (carbon-based and Pb stylus) to sketch the figure and then refined the details and shadows with iron-gall ink. The paper presents a methodological approach for the analytical examination of fragile paper artworks.

Scanning multispectral IR reflectography SMIRR: an advanced tool for art diagnostics.

Spectral imaging technology, widely used in remote sensing applications, such as satellite or radar imaging, has recently gained importance in the field of artwork conservation. In particular, multispectral imaging in the near-infrared region (NIR) has proved useful in analyzing ancient paintings because of the transparency of most pigments and their varied reflectance changes over this spectral region. A variety of systems, with different detectors and filtering or dispersing technologies, have been implemented. Despite the recognized potential of multispectral NIR imaging, which provides information on both spectral and spatial domains (thus extending the capabilities of conventional imaging and spectroscopy), most of the systems currently used in art diagnostics have limitations. The technology is still in its early stages of development in this field. In this Account, we present the scanning multispectral IR reflectography (SMIRR) technique for artwork analysis, together with an integrated device for the acquisition of imaging data. The instrument prototype is a no-contact optical scanner with a single-point measurement of the reflectance, capable of simultaneously collecting a set of 14 spatially registered images at different wavelengths in the NIR range of 800-2300 nm. The data can be analyzed as a spectral cube, both as a stack of wavelength resolved images (multi-NIR reflectography) and as a series of point reflectance spectra, one for each sampled pixel on the surface (NIR spectrometry). We explore the potential of SMIRR in the analysis of ancient paintings and show its advantages over the wide-band conventional method. The multispectral option allows the choice of the most effective NIR bands and improves the ability to detect hidden features. The interband comparison aids in localizing areas of different pictorial materials with particular NIR reflectance. In addition to the analysis of single monochromatic images, enhancement procedures involving the joint processing of multispectral planes, such as subtraction and ratio methods, false color representation, and statistical tools such as principal component analysis, are applied to the registered image dataset for extracting additional information. Maintaining a visual approach in the data analysis allows this tool to be used by museum staff, the actual end-users. We also present some applications of the technique to the study of Italian masterpieces, discussing interesting preliminary results. The spectral sensitivity of the detection system, the quality of focusing and uniformity of the acquired images, and the possibility for selective imaging in NIR bands in a registered dataset make SMIRR an exceptional tool for nondestructive inspection of painting surfaces. The high quality and detail of SMIRR data underscore the potential for further development in this field.