Dual mode standoff imaging spectroscopy documents the painting process of the Lamb of God in the Ghent Altarpiece by J. and H. Van Eyck.

The ongoing conservation treatment program of the Ghent Altarpiece by Hubert and Jan Van Eyck, one of the iconic paintings of the west, has revealed that the designs of the paintings were changed several times, first by the original artists, and then during later restorations. The central motif, The Lamb of God, representing Christ, plays an essential iconographic role, and its depiction is important. Because of the prevalence of lead white, it was not possible to visualize the Van Eycks' original underdrawing of the Lamb, their design changes, and the overpaint by later restorers with a single spectral imaging modality. However, by using elemental (x-ray fluorescence) and molecular (infrared reflectance) imaging spectroscopies, followed by analysis of the resulting data cubes, the necessary chemical contrast could be achieved. In this way, the two complementary modalities provided a more complete picture of the development and changes made to the Lamb.

Molecular Fluorescence Imaging Spectroscopy for Mapping Low Concentrations of Red Lake Pigments: Van†Gogh's Painting The Olive Orchard.

Vincent van Gogh used fugitive red lake pigments that have faded in some paintings. Mapping their distribution is key to understanding how his paintings have changed with time. While red lake pigments can be identified from microsamples, in situ identification and mapping remain challenging. This paper explores the ability of molecular fluorescence imaging spectroscopy to identify and, more importantly, map residual non-degraded red lakes. The high sensitivity of this method enabled identification of the emission spectra of eosin (tetrabromine fluorescein) lake mixed with lead or zinc white at lower concentrations than elemental X-ray fluorescence (XRF) spectroscopy used on account of bromine. The molecular fluorescence mapping of residual eosin and two carmine red lakes in van Gogh's The Olive Orchard is demonstrated and compared with XRF imaging spectroscopy. The red lakes are consistent with the composition of paint tubes known to have been used by van Gogh.

Standoff Mid-Infrared Emissive Imaging Spectroscopy for Identification and Mapping of Materials in Polychrome Objects.

Microscale mid-infrared (mid-IR) imaging spectroscopy is used for the mapping of chemical functional groups. The extension to macroscale imaging requires that either the mid-IR radiation reflected off or that emitted by the object be greater than the radiation from the thermal background. Reflectance spectra can be obtained using an active IR source to increase the amount of radiation reflected off the object, but rapid heating of greater than 4 °C can occur, which is a problem for paintings. Rather than using an active source, by placing a highly reflective tube between the painting and camera and introducing a low temperature source, thermal radiation from the room can be reduced, allowing the IR radiation emitted by the painting to dominate. Thus, emissivity spectra of the object can be recovered. Using this technique, mid-IR emissivity image cubes of paintings were collected at high collection rates with a low-noise, line-scanning imaging spectrometer, which allowed pigments and paint binders to be identified and mapped.

Macroscale multimodal imaging reveals ancient painting production technology and the vogue in Greco-Roman Egypt.

Macroscale multimodal chemical imaging combining hyperspectral diffuse reflectance (400-2500 nm), luminescence (400-1000 nm), and X-ray fluorescence (XRF, 2 to 25 keV) data, is uniquely equipped for noninvasive characterization of heterogeneous complex systems such as paintings. Here we present the first application of multimodal chemical imaging to analyze the production technology of an 1,800-year-old painting and one of the oldest surviving encaustic ("burned in") paintings in the world. Co-registration of the data cubes from these three hyperspectral imaging modalities enabled the comparison of reflectance, luminescence, and XRF spectra at each pixel in the image for the entire painting. By comparing the molecular and elemental spectral signatures at each pixel, this fusion of the data allowed for a more thorough identification and mapping of the painting's constituent organic and inorganic materials, revealing key information on the selection of raw materials, production sequence and the fashion aesthetics and chemical arts practiced in Egypt in the second century AD.

Complementary standoff chemical imaging to map and identify artist materials in an early Italian Renaissance panel painting.

Two imaging modalities based on molecular and elemental spectroscopy were used to characterize a painting by Cosimo Tura. Visible-to-near-infrared (400-1680 nm) reflectance imaging spectroscopy (RIS) and X-ray fluorescence (XRF) imaging spectroscopy were employed to identify pigments and determine their spatial distribution with higher confidence than from either technique alone. For example, Mary's red robe was modeled through the distribution of an insect-derived red lake (RIS map) and lead white (XRF lead map), rather than a layer of red lake on vermilion. The RIS image cube was also used to isolate the preparatory design by mapping the reflectance spectra associated with it. In conjunction with results from an earlier RIS study (1650-2500 nm) to map and identify the binding media, a more thorough understanding was gained of the materials and techniques used in the painting.

Mapping of egg yolk and animal skin glue paint binders in Early Renaissance paintings using near infrared reflectance imaging spectroscopy.

In situ chemical imaging techniques are being developed to provide information on the spatial distribution of artists' pigments used in polychrome works of art such as paintings. The new methods include reflectance imaging spectroscopy and X-ray fluorescence mapping. Results from these new methods have extended the knowledge obtained from site-specific chemical analyses widely in use. While these mapping methods have aided in determining the distribution of pigments, there is a growing interest to develop methods capable of identifying and mapping organic paint binders as well. Near infrared (NIR) reflectance spectroscopy has been extensively used in the remote sensing field as well as in the chemical industry to detect organic compounds. NIR spectroscopy provides a rapid method to assay organics by utilizing vibrational overtones and combination bands of fundamental absorptions that occur in the mid-IR. Here we explore the utility of NIR reflectance imaging spectroscopy to map organic binders in situ by examining a series of panel paintings known to have been painted using distemper (animal skin glue) and tempera (egg yolk) binders as determined by amino acid analysis of samples taken from multiple sites on the panels. In this report we demonstrate the success in identifying and mapping these binders by NIR reflectance imaging spectroscopy in situ. Three of the four panel paintings from Cosimo Tura's The Annunciation with Saint Francis and Saint Louis of Toulouse (ca. 1475) are imaged using a highly sensitive, line-scanning hyperspectral imaging camera. The results show an animal skin glue binder was used for the blue skies and blue robe of the Virgin Mary, and egg yolk tempera was used for the red robes and brown landscape. The mapping results show evidence for the use of both egg yolk and animal skin glue in the faces of the figures. The strongest absorption associated with lipidic egg yolk features visually correlates with areas that appear to have white highlights. The results are in agreement with prior site-specific amino acid analysis, underscoring the synergy of both methods. The work here demonstrates that NIR reflectance imaging spectroscopy is a useful technique that can identify and map paint binding media based on differences in chemical composition.

Stress mapping of undamaged, strained, and failed regions of bone using Raman spectroscopy.

Stress differences via spectral shifts that arise among failed, strained, and undamaged regions of bone can be determined using Raman spectroscopy and double-notch specimens. A double-notch specimen is a model in which the early stages of fracture can be examined. At four-point bending, fracture occurs at one of the notches. Tissue near each notch is representative of bone in a state either directly before or after bone failure. Raman images are acquired among three regions: control, strained (root of unbroken notch), and failed (root of fractured notch). The center of gravities (CGs), a way to monitor wavenumber shifts, of the phosphate v(1) band are calculated. A PO(4) (-3) v(1) band shift most likely corresponds to a change in spacing between phosphate cations and anions. This spectral shift is converted into stress values using the dvdP coefficient, determined by applying known pressures/stresses and measuring the change in position of the PO(4) (-3) v(1) band. In comparison to control regions, the residual stress in strained and failed regions is significantly higher (p=0.0425 and p=0.0169, respectively). In strained regions, residual stress is concentrated near the corners of the unbroken notch, whereas in failed regions the high stresses are confined near the edge of the fracture.

Transcutaneous Raman spectroscopy of murine bone in vivo.

Raman spectroscopy can provide valuable information about bone tissue composition in studies of bone development, biomechanics, and health. In order to study the Raman spectra of bone in vivo, instrumentation that enhances the recovery of subsurface spectra must be developed and validated. Five fiber-optic probe configurations were considered for transcutaneous bone Raman spectroscopy of small animals. Measurements were obtained from the tibia of sacrificed mice, and the bone Raman signal was recovered for each probe configuration. The configuration with the optimal combination of bone signal intensity, signal variance, and power distribution was then evaluated under in vivo conditions. Multiple in vivo transcutaneous measurements were obtained from the left tibia of 32 anesthetized mice. After collecting the transcutaneous Raman signal, exposed bone measurements were collected and used as a validation reference. Multivariate analysis was used to recover bone spectra from transcutaneous measurements. To assess the validity of the transcutaneous bone measurements cross-correlations were calculated between standardized spectra from the recovered bone signal and the exposed bone measurements. Additionally, the carbonate-to-phosphate height ratios of the recovered bone signals were compared to the reference exposed bone measurements. The mean cross-correlation coefficient between the recovered and exposed measurements was 0.96, and the carbonate-to-phosphate ratios did not differ significantly between the two sets of spectra (p > 0.05). During these first systematic in vivo Raman measurements, we discovered that probe alignment and animal coat color influenced the results and thus should be considered in future probe and study designs. Nevertheless, our noninvasive Raman spectroscopic probe accurately assessed bone tissue composition through the skin in live mice.

Image-guided Raman spectroscopic recovery of canine cortical bone contrast in situ.

Raman scattering provides valuable biochemical and molecular markers for studying bone tissue composition with use in predicting fracture risk in osteoporosis. Raman tomography can image through a few centimeters of tissue but is limited by low spatial resolution. X-ray computed tomography (CT) imaging can provide high-resolution image-guidance of the Raman spectroscopic characterization, which enhances the quantitative recovery of the Raman signals, and this technique provides additional information to standard imaging methods. This hypothesis was tested in data measured from Teflon tissue phantoms and from a canine limb. Image-guided Raman spectroscopy (IG-RS) of the canine limb using CT images of the tissue to guide the recovery recovered a contrast of 145:1 between the cortical bone and background. Considerably less contrast was found without the CT image to guide recovery. This study presents the first known IG-RS results from tissue and indicates that intrinsically high contrasts (on the order of a hundred fold) are available.

Noninvasive Raman tomographic imaging of canine bone tissue.

Raman spectroscopic diffuse tomographic imaging has been demonstrated for the first time. It provides a noninvasive, label-free modality to image the chemical composition of human and animal tissue and other turbid media. This technique has been applied to image the composition of bone tissue within an intact section of a canine limb. Spatially distributed 785-nm laser excitation was employed to prevent thermal damage to the tissue. Diffuse emission tomography reconstruction was used, and the location that was recovered has been confirmed by micro-computed tomography (micro-CT) images.

Optical clearing in transcutaneous Raman spectroscopy of murine cortical bone tissue.

The effect of optical clearing with glycerol on the Raman spectra of bone tissue acquired transcutaneously on right and left tibiae from four mice is studied. Multiple transcutaneous measurements are obtained from each limb; glycerol is then applied as an optical clearing agent, and additional transcutaneous measurements are taken. Glycerol reduces the noise in the raw spectra (p=0.0037) and significantly improves the cross-correlation between the recovered bone factor and the exposed bone measurement in a low signal-to-noise region of the bone spectra (p=0.0245).

Subsurface and transcutaneous Raman spectroscopy and mapping using concentric illumination rings and collection with a circular fiber-optic array.

Different spatial separations between an illumination ring and a bundle of 50 collection fibers focused to collect light in the center of the ring were used to investigate the recovery of subsurface Raman spectra. The depth of Raman signal recovery and the preservation of spatial information in the recovered signal were investigated using polymer blocks stacked in different geometries. The illumination rings were then combined into a single data set to increase variation in the signal. Multivariate data analysis was used to recover the Raman spectra of the subsurface component. The Raman spectrum of a Delrin target was recoverable at depths up to 22.6 mm of overlying Teflon. Spatial information was lost at approximately 6.5 mm below the Teflon surface. The same protocols were used to recover canine bone spectra transcutaneously at depths up to 5 mm below the skin's surface. The recovered bone spectra were validated by exposed bone measurements.

Transcutaneous fiber optic Raman spectroscopy of bone using annular illumination and a circular array of collection fibers.

Transcutaneous bone Raman spectroscopy with an exciting annulus of 785-nm laser light surrounding the field of view of a circular array of collection fibers is demonstrated. The configuration provides distributed laser light. The annulus is located 2 to 3 mm beyond the edge of the field of view of the collection fibers to reject contributions from skin and other overlying tissues. Data are presented for rat and chicken tissue. For rat tibia, the carbonate/phosphate ratio measured at a depth of 1 mm below the skin is in error by 2.3% at an integration time of 120 s and within 10% at a 30-s integration time. For chicken tibia 4 mm below the skin surface, the error is less than 8% with a 120-s integration time.