Periodic Reporting for period 1 - SCIMITAR (Synergistic Combination of Immunolabeling and Molecular Imaging Technologies for Art Research)
Periodo di rendicontazione: 2023-10-23 al 2025-10-22
This project falls within the scope of initiatives aimed at understanding and preserving Cultural Heritage, so that it can be better passed on and preserved for future generations. When focusing on an art object, it is not only essential to study its structure, creation, history, and significance, but it is also necessary to understand its past and current material state as well as projected changes or alterations in the future. This is a continuous and complex process which requires deep interdisciplinary collaboration. In the case of historical paintings, which are heterogeneous objects, made of stacked thin layers containing both organic and inorganic ingredients that have dried together, aged and interacted within uncontrolled environments over long periods of time, one of the multiple approaches to increasing our knowledge of them is to study how macroscopic observations correlate with chemical make-up at the microscopic level. The aim of SCIMITAR was to take part in these efforts using mass spectrometry imaging, and especially Time-of-Flight Secondary Ion Mass Spectrometry (ToF-SIMS), to investigate the microstructure of the dried paint. These chemical imaging techniques have grown in relevance in the field of heritage science over the last decades, as they are especially suited for heterogeneous systems. ToF-SIMS can for instance simultaneously localize and identify both organic and mineral components (such as pigments, binders, degradation products), and its sub-micrometer spatial resolution is very valuable to investigate painting micro-samples that consist of sub-millimeter stacked layers of dried paint that contain both organic and inorganic ingredients. While studies over the last two decades have attested to the potential of ToF-SIMS in providing information relevant for investigating painting samples, several challenges must be addressed to fully integrate ToF-SIMS imaging into the heritage research toolbox.
ToF-SIMS imaging yields multidimensional, complex datasets where much of the information is not fully understood and therefore often with a large proportion remaining unexploited, or not accessible due to technical limitations. As an example, the high energy ion source causes large biomolecules such as proteins to fragment, making it impossible to identify them intact. As a variety of protein-rich ingredients coexist and age in paintings, being able to identify them while accessing their spatial distribution at the micrometer scale would be extremely valuable. The project aims at developing methods to reduce the ambiguity of the materials identified in the heterogeneous microstructures of old paintings samples using a ToF-SIMS instrument, by addressing specific analytical limitations. Correlation between materials and their signal can be investigated using novel Tandem MS imaging capabilities and a well-documented material library from conservation institutions. Additionally, the project aims at developing alternative ways to allow targeted imaging of proteins in painting materials using ToF-SIMS, for instance thanks to its combination with immunohistochemistry relying on metal-labeled antibodies. The third aspect addressed within the aim of this project is to improve the shareability and accessibility of the technique by facilitating the data processing, by sharing databases, and by communicating the capabilities and limitation of the techniques to enable researchers from all disciplines to independently evaluate if the performance achievable using ToF-SIMS imaging can answer specific research questions relevant to a given sample.
At the end of the project, dissemination of the collected data, established protocols, and developed methodologies will advance the mass spectrometry imaging field toward achieving routine ToF-SIMS imaging of cultural heritage materials.
ToF-SIMS imaging yields multidimensional, complex datasets where much of the information is not fully understood and therefore often with a large proportion remaining unexploited, or not accessible due to technical limitations. As an example, the high energy ion source causes large biomolecules such as proteins to fragment, making it impossible to identify them intact. As a variety of protein-rich ingredients coexist and age in paintings, being able to identify them while accessing their spatial distribution at the micrometer scale would be extremely valuable. The project aims at developing methods to reduce the ambiguity of the materials identified in the heterogeneous microstructures of old paintings samples using a ToF-SIMS instrument, by addressing specific analytical limitations. Correlation between materials and their signal can be investigated using novel Tandem MS imaging capabilities and a well-documented material library from conservation institutions. Additionally, the project aims at developing alternative ways to allow targeted imaging of proteins in painting materials using ToF-SIMS, for instance thanks to its combination with immunohistochemistry relying on metal-labeled antibodies. The third aspect addressed within the aim of this project is to improve the shareability and accessibility of the technique by facilitating the data processing, by sharing databases, and by communicating the capabilities and limitation of the techniques to enable researchers from all disciplines to independently evaluate if the performance achievable using ToF-SIMS imaging can answer specific research questions relevant to a given sample.
At the end of the project, dissemination of the collected data, established protocols, and developed methodologies will advance the mass spectrometry imaging field toward achieving routine ToF-SIMS imaging of cultural heritage materials.
The project is divided into 3 work packages (WP) that can be summarized as follows:
WP1: Improve methodology and expand the database for high-resolution imaging mass spectrometry applied to historical painting materials.
Selection of reference materials, sacrificial historical samples, and historical samples in concertation with the partners of the project. Preparation of mock-ups with controlled structure and composition to answer specific analytical questions and assess the exploitability of ToF-SIMS signal in identified ambiguous cases (mixtures of organics, interface between layers).
Analysis of the accessible large set of samples to enlarge a general database, then data processing and comparison to identify reliable ion markers for specific materials to be shared with the CH community
Characterization with high resolution ToF-SIMS imaging of historical samples on which specific research questions were formulated, then assessment of the relevance of the result in regard with the research question in concertation with the expertise of partners providing the sample.
Development of a sample preparation protocol dedicated to improving the data quality for historical cross-section when using ToF-SIMS (reduction of the detrimental effect of the insulating embedding material)
Selection of discriminating markers to be explored using tandem MS
Samples documented and kept in the host laboratories to enable further studies involving the effect of ageing.
List of reliable marker ions tracing back to pigments have been published within the time of the project
WP2: Identification and spatial distribution of proteins
Definition of target proteins thanks to the comparison of datasets from proteinic materials from WP1 (yolk vitellogenin, albumin, collagen, lipoproteins)
Improved understanding of what ion can relate to protein detection in a regular ToF-SIMS dataset by comparison with other MSI instruments (MALDI, LDI) on specifically designed samples during WP1
Identification of fragments specific to the combination of egg yolk / copper pigment and egg yolk / lead pigments that can serve as internal label in painting samples for protein. Assessment of their detectability in historical sample cross sections.
WP3: ToF-SIMS data representation software development and sharing
Interface development: definition of the specifications answering the needs evaluated by discussing with partners / collaborators during the project
Construction of the general database needed to attribute an ion to a signal in the interface
Edition of general guide providing method for analysis, data processing and a tutorial to the interface
WP1: Improve methodology and expand the database for high-resolution imaging mass spectrometry applied to historical painting materials.
Selection of reference materials, sacrificial historical samples, and historical samples in concertation with the partners of the project. Preparation of mock-ups with controlled structure and composition to answer specific analytical questions and assess the exploitability of ToF-SIMS signal in identified ambiguous cases (mixtures of organics, interface between layers).
Analysis of the accessible large set of samples to enlarge a general database, then data processing and comparison to identify reliable ion markers for specific materials to be shared with the CH community
Characterization with high resolution ToF-SIMS imaging of historical samples on which specific research questions were formulated, then assessment of the relevance of the result in regard with the research question in concertation with the expertise of partners providing the sample.
Development of a sample preparation protocol dedicated to improving the data quality for historical cross-section when using ToF-SIMS (reduction of the detrimental effect of the insulating embedding material)
Selection of discriminating markers to be explored using tandem MS
Samples documented and kept in the host laboratories to enable further studies involving the effect of ageing.
List of reliable marker ions tracing back to pigments have been published within the time of the project
WP2: Identification and spatial distribution of proteins
Definition of target proteins thanks to the comparison of datasets from proteinic materials from WP1 (yolk vitellogenin, albumin, collagen, lipoproteins)
Improved understanding of what ion can relate to protein detection in a regular ToF-SIMS dataset by comparison with other MSI instruments (MALDI, LDI) on specifically designed samples during WP1
Identification of fragments specific to the combination of egg yolk / copper pigment and egg yolk / lead pigments that can serve as internal label in painting samples for protein. Assessment of their detectability in historical sample cross sections.
WP3: ToF-SIMS data representation software development and sharing
Interface development: definition of the specifications answering the needs evaluated by discussing with partners / collaborators during the project
Construction of the general database needed to attribute an ion to a signal in the interface
Edition of general guide providing method for analysis, data processing and a tutorial to the interface
The accessibility and relevance of ToF-SIMS imaging in Cultural Heritage research is still limited, while there is a growing existing knowledge regarding what the technique can provide when investigating historical painting samples, or more generally any heterogeneous micro-sample from Cultural Heritage object. One of the main assets of the technique is that, given that a sample has been taken (for another study for instance), performing ToF-SIMS imaging will not impact the integrity of the sample. Technically, it could be applied to any cross-section stored in sample libraries from conservation institutions after past studies, even after decades. The difficulty of accessing an instrument, an operator, and avoiding time-consuming data processing are the main reasons preventing this from happening.
The SCIMITAR project dedicated research time fully to tackle some of the existing analytical challenges to widen its impact in the interdisciplinary context of investigating heritage materials. By defining and sharing reliable markers confidently tracing back to a given compound, SCIMITAR fosters the processing of new datasets of Cultural Heritage samples but also helps the analysis of disregarded existing datasets. The sample preparation and analysis protocol could be compared, adjusted and improved on a large range of historical cross-sections to define an approach ensuring meaningful reproducible signal, and to evaluate the impact of the sample features (thickness of the block, layer dimensions) on the technical limitations. This enables to provide projected capabilities of the technique in the early stage of collaborations, to refine research questions. Additionally, sharing information and transferring knowledge with the broader cultural heritage research community is essential to advance from isolated case studies to more systematic interdisciplinary ToF-SIMS application.
These main outcomes of the project will be incorporated in an open resource destined to Cultural Heritage community, including the general spectral database, a data processing/browsing tool compatible with both new and existing datasets, and an online guide compiling all the existing knowledge on sample preparation, data acquisition, data processing and interpretation specifically dedicated to painting materials.
The SCIMITAR project dedicated research time fully to tackle some of the existing analytical challenges to widen its impact in the interdisciplinary context of investigating heritage materials. By defining and sharing reliable markers confidently tracing back to a given compound, SCIMITAR fosters the processing of new datasets of Cultural Heritage samples but also helps the analysis of disregarded existing datasets. The sample preparation and analysis protocol could be compared, adjusted and improved on a large range of historical cross-sections to define an approach ensuring meaningful reproducible signal, and to evaluate the impact of the sample features (thickness of the block, layer dimensions) on the technical limitations. This enables to provide projected capabilities of the technique in the early stage of collaborations, to refine research questions. Additionally, sharing information and transferring knowledge with the broader cultural heritage research community is essential to advance from isolated case studies to more systematic interdisciplinary ToF-SIMS application.
These main outcomes of the project will be incorporated in an open resource destined to Cultural Heritage community, including the general spectral database, a data processing/browsing tool compatible with both new and existing datasets, and an online guide compiling all the existing knowledge on sample preparation, data acquisition, data processing and interpretation specifically dedicated to painting materials.