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The extraction and identification of silk residues in tombs is of great significance for studying the distribution and spread of early silk. However, the complex organic matter in the tomb hinders the accurate identification of silk. In this study, a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) based on immunomagnetic beads (IMBs) was developed for the rapid enrichment and detection of silk residues. The double-antibody sandwich ELISA method established by pairing the IMBs prepared by the silk fibroin monoclonal antibody SF-3 and the silk fibroin monoclonal-labeled antibody bio-SF-1 had the highest detection sensitivity, with a linear detection range of 10 to 10 4 ng mL -1 and a detection limit of 5.12 ng mL -1 . This method was excellent in the extraction and analysis of silk residues from archaeological imprints and soil samples and successfully identified silk residues in samples at the final stage of silk degradation (physical invisible silk). The proteomics analysis results demonstrated the feasibility and practicability of this method.

The process and mechanism of silk degradation is still a bewildering mystery in the investigation and conservation of cultural relics, which rely on the development of accurate and tailored analysis technologies. Here, two advanced approaches, proteomics and immunology, were developed for determining the deterioration behavior of historic silk fabrics and artificially aged samples from the molecular to the holistic level. The surface morphology and secondary structure of silk were destroyed during degradation. Subsequently, the proteomics and immunology analysis demonstrated a new degradation model differing from previous reports. First, the amorphous region and the looser crystalline regions were destroyed together, and the macromolecular chains were broken randomly. Then, the tight β-sheet blocks in the crystalline region were exposed and deteriorated, which expedited the degradation of tight β-sheet blocks and relatively loose blocks in the crystalline domain as well as the amorphous domain, ultimately yielding small molecule polypeptides. However, the deterioration process of ancient fabrics could be accelerated by poor burial conditions, thus showing distinct destructive characteristics. Overall, the results gave us a more comprehensive and profound understanding of the degradation process of ancient silk.

The ultrasensitive detection of fibroin in unearthed silk relics has great significance for investigating the origin and transmission of silk. In this study, an anti-fibroin monoclonal antibody was successfully prepared through animal immunization. Next, a label-free electrochemical immunosensor was fabricated using layer-by-layer self-assembly technology, and an indirect enzyme-linked immunosorbent assay (ELISA) was proposed. The two methods exhibited excellent sensitivity and specificity in the detection of silk fibroin, while the immunosensor showed a wider quantitative detection range (0.1-100 ng mL -1 ) and a lower detection limit (0.051 ng mL -1 ) than ELISA (10-100 ng mL -1 and 8.71 ng mL -1 ). Furthermore, the performance of the immunosensor was superior in archaeological sample detection. Taking advantage of the well-prepared monoclonal antibody, the two proposed immunological assays demonstrate tremendous potential for the ultrasensitive detection of silk fibroin, which can make great contributions to exploring the origin and transmission routes of ancient silks., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Achieving efficient identification of silk protein requires highly sensitive analytical techniques and favorable extraction methods, which is of great significance to the research of ancient silk, especially for the controversial issue of the silk origin. In this paper, proteomics and western blot were proposed to analyze the silk proteins of Bombyx mori (B. mori) and Antheraea pernyi (A. pernyi) dissolved by different methods. First, the differences in secondary structure were detected via spectroscopy. LC-MS/MS was then employed to characterize the peptides of silk proteins precisely. LiBr solution exhibited outstanding dissolution effect on B. mori cocoon, with 87 proteins detected; while copper-ethylenediamine solution (CED) was more appropriate for A. pernyi cocoon, and 16 proteins were identified in A. pernyi-CED. In addition to fibroin and sericin, abundant seroins, enzymes, protease inhibitors, other functional proteins and uncharacterized proteins were detected. Based on the LC-MS/MS data, diagnostic antibodies for the two species were prepared, and fibroin was successfully identified by western blot assay because both dissolution methods were gentle and did not destroy the antigenic epitopes in the protein molecule. Owing to their good specificity and high sensitivity, these diagnostic antibodies have good application prospects in immunoassays of different silk species. SIGNIFICANCE: This study presents the comprehensive analysis on silk identification of proteins from B. mori and A. pernyi extracted by different methods via the proteomic and immunology as well as the conventional approaches. Great coverage of two cocoon proteomes was accomplished, which demonstrated the outstanding difference in components and abundance. Based on the proteomics analysis, the diagnostic antibodies against two species were prepared and identified the corresponding fibroin successfully in the completed protein mixtures. To our knowledge, the proteomic and immunology procedures with high efficiency, sensitivity and specificity are novel analysis on the silk identification and has great potential in the field of ancient silk detection., (Copyright © 2019. Published by Elsevier B.V.)

In recent years, increasing attention has been paid to the origin, transmission and communication of silk. However, this is still an unsolved mystery in archaeology. The identification of silk-producing species, especially silk produced by Bombyx mori (B. mori) and Antheraea pernyi (A. pernyi), is of key significance to address this challenge. In this study, two innovative methods, i.e. immunology and proteomics, were proposed and successfully established for the species identification of silks. ELISAs result demonstrated that the two prepared antibodies exhibited high sensitivity and specificity in distinguishing B. mori and A. pernyi silk. No cross-reactivity with each other was observed. Moreover, biomarkers were obtained for Bombyx and Antheraea through proteomic analysis. It was also confirmed that the biomarkers were suitable for identifying the species that produced a given silk sample. Compared with conventional methods for distinguishing silk species, immunological and proteomics techniques used in tandem can provide intact information and have the potential to provide accurate and reliable information for species identification.

Species identification is of key significance for exploring the origin and transmission of ancient silks. In this study, two novel methods, i.e. western blot (WB) and proteomics analyses, were proposed and established to identify the differences between silks from Bombyx mori (B. mori) and two other distinctive species (Eri silkworm and Chestnut silkworm). Three diagnostic antibodies, a polyclonal anti-silk fibroin (anti-SF) antibody (pAb), a polyclonal anti-SF-specific peptide antibody (pAsb), and a monoclonal anti-SF antibody (mAb) were designed and prepared to distinguish silk species using the antibody-based WB technique. Proteomics analysis by liquid chromatography-tandem mass spectrometry was performed to further identify silk species at the protein level. WB results indicated that the three antibodies showed high specificity and affinity and could discern B. mori silk from Eri and Chestnut silks. Biomarkers for each SF were obtained using proteomics analysis, and they have the potential to serve as standards for identifying silk species. Thus, combining WB and proteomics analyses with conventional methods can provide more accurate silk information and may be suitable for identifying other proteinaceous materials in archaeological field.

Recently, much interest has been paid to the separation of silk produced by Bombyx mori from silk produced by other species and tracing the beginnings of silk cultivation from wild silk exploitation. In this paper, significant differences between silks from Bombyx mori and other species were found by microscopy and spectroscopy, such as morphology, secondary structure, and amino acid composition. For further accurate identification, a diagnostic antibody was designed by comparing the peptide sequences of silks produced by Bombyx mori and other species. The results of the noncompetitive indirect enzyme-linked immunosorbent assay (ELISA) indicated that the antibody that showed good sensitivity and high specificity can definitely discern silk produced by Bombyx mori from silk produced by wild species. Thus, the antibody-based immunoassay has the potential to be a powerful tool for tracing the beginnings of silk cultivation. In addition, combining the sensitive, specific, and convenient ELISA technology with other conventional methods can provide more in-depth and accurate information for species identification.

This work studied the concretion components of ancient silk fabrics unearthed from Nanjing, China. Based on the analysis of the components, mechanism of concretion of the silk fabrics were proposed. Physical, chemical, and biological factors were considered as possible reasons for the concretion of ancient silk fabrics. Mechanical extrusion or falling, crystal bonding, Van der walls forces, hydrogen bonding, covalent bonding, and ionic bonding are purposed as possible influences and/or interactions between the concretion components and silk fabrics. The concretion mechanism, which includes common mineralization nucleation, re-crystallization, bio-mineralization nucleation and crystallization are discussed separately. Mineralization nucleation and crystallization have a great influence on the aging, hardening (even rotting), and final concreting of buried silk fabrics. Possible processing models were established according to the results obtained. This study presents new viewpoints to understand the concretion of unearthed silk fabrics, which could provide theoretical evidence for the unfolding of concreted silk fabrics, especially those fragile, even rotten ones. [ABSTRACT FROM AUTHOR]

The stable isotope technique provides the possibility to trace ancient textiles because the technique is associated with advantages such as trace indication, fast detection, and accurate results. Since different cocooning conditions may impact cocoons even under identical habitats, it is important to investigate the effects of different cocooning temperatures and humidity on the isotope incorporation values in the cocoons. In this study, silk fibers were reeled under different conditions of temperature and humidity, followed by analysis of the secondary structure of cocoon proteins and isotope incorporation patterns. We found that the deviations in carbon isotope values of silk under different cocooning conditions could reach up to 0.76‰, while the deviation in carbon isotope values at different locations of a single silk was 2.75‰. Further, during the cocooning process, depletion of the 13C-isotope at different locations of the silk fibers was observed, reducing the δ13C values. We proposed that the changes in carbon isotopes in silk were related to the content of sericin and silk fibroin in silk. Finally, we did not observe a significant difference in isotope ratios in degummed cocoons. In summary, the 13C isotope was enriched in sericin, whereas 15N was enriched in fibroin, and these findings provide basic information for tracing the provenance of silks. [ABSTRACT FROM AUTHOR]

The characterization of ancient silk provides significant historical information and contribute to the identification of the origin of the material. At present, there are many ways to characterize silk. However, on-site identification of ancient silk remains a crucial challenge. In this study, a paper chromatography system consisting of a colloidal gold immunochromatographic strip (GICS) and a time-resolved fluorescence immunochromatographic strip (TICS) is proposed to meet this challenge. On the basis of the original paper chromatography technique, a monoclonal antibody against silk fibroin (SF) was prepared and applied in this new paper chromatography system. Both the GICS and the TICS showed good sensitivity, specificity and reliability, with the limit of detection for silk fibroin being 30 μg/mL and 49.20 ng/mL, respectively. Moreover, there was no cross reaction with possible interferents. Furthermore, the GICS had acceptable thermal stability. The recovery of the TICS assay was in the range from 98.12% to 103%, and the variation coefficient was from 5.34% to 6.93%. This work significantly improves access to scientific information at archaeological sites, providing new scientific evidence for exploring the origin and transmission roadmap of ancient silk. [ABSTRACT FROM AUTHOR]

The existing historical silk, which has great historical and academic research value, is an indispensable and important carrier for the study of global ancient civilization. It is very important to scientifically evaluate the preservation status of such silk. In this paper, the corresponding relationship between the surface resistance and the retention rate of breaking strength was established by testing simulated samples of photoaged silk fabrics. The effectiveness of the surface resistance method was suggested by amino acid analysis and infrared spectroscopic analysis. The test results show that the surface resistance method is a simple and fast nondestructive test to establish improved rules of consideration for the aging degree of silk relics. According to the corresponding relationship between the surface resistance and the retention rate of breaking strength, the current breaking strength of the fibers of historical silk can be inferred, which provides a scientific basis for the aging evaluation and protection of historical silk. [ABSTRACT FROM AUTHOR]

Mineralized textiles often unearthed along with metal objects, are textile residues in which the fibers are partially or completely replaced by minerals. Some textile fragments with intact fiber shapes but mineralization on the surface of a ring-shaped iron sword, had recently been found from Maoling Mausoleum in Shanxi province, China. In this study, three-dimension microscope, scanning electron microscope-energy disperse spectroscopy (SEM-EDS) and Raman spectroscopy were used to observe the morphology of fiber and analyze the mineral element information, and two immunology testing techniques would be used to determine whether the textile fragments included the silk protein. The results showed that the hilt string imprint (sample A) was a twill braiding with cinnabar remaining on the surface and the blade textile imprint (sample B) was a tabby fabric, and these fibers were mineralized by iron ions to form a cavity structure. The morphology of sample B was similar to that of mulberry silk with a smooth surface and a triangular cross section. At the same time, silk protein was successfully detected in a few milligrams of sample B using fibroin polyclonal antibody by Enzyme-linked Immunosorbent assay (ELISA) and immunofluorescence method (IFM). Thus, it can be inferred that the iron sword was covered or wrapped in silk at that time. This research provided a technical support for the identification of silk residues in mineralized textiles, which is of great significance for the study of early organic fabrics. • Mineralized samples were found on the surface of a ring-shaped iron sword. • Morphologies of mineralized fabrics were observed with microscopic techniques. • Silk protein was detected in mineralized sample using two immunological methods. [ABSTRACT FROM AUTHOR]

The origin and dissemination of silk have been hotly debated in the field of archaeology, and the key to resolving this controversy lies in the detection and species identification of ancient silk microtraces. Herein, a taxonomically specific anti-fibroin monoclonal antibody was successfully prepared and a layer-by-layer self assembly electrochemical immunosensor was innovatively proposed for detecting silk traces based on flexible carbon cloth. The immunosensor possessed a broad linear range of 10−2-103 ng mL−1 and a detection limit of 2.15 pg mL−1 for the ultrasensitive detection of Bombyx mori silk traces. In addition, the elaborate immunosensor exhibited satisfactory high specificity, storage stability and reproducibility. In particular, the qualitative and quantitative performance of the immunosensor was excellent in the analysis of archaeological samples. Therefore, this work demonstrates that the proposed method not only provides a reliable analytical tool for exploring the origin and spread of archaeological silk, but also improves our understanding of how to use emerging materials like two-dimensional titanium carbide to creat innovative biosensors. [ABSTRACT FROM AUTHOR]

• Stable isotope technique was firstly used to analyze ancient and artificial aged silk fabrics. • The stable isotope ratios of ancient silk fabrics under various excavation environments display different trends. • Heavy stable isotopes were enriched during degradation of the silk fabrics under most aging conditions. • Degradation of silk fabrics is a significant cause of stable isotope offset in historical silk fabrics. • Different aging factors should be considered when tracing the origin of historic silk fabrics using stable isotope techniques. Understanding the degradation of silk fibers and tracing silk fabrics from archeological sites have always been challenging. The stable isotope technique provides the possibility to trace ancient textiles, especially for the controversial issue of the silk origin. Chinese historic silk fabrics from the Xia dynasty (about 4000 years ago), Warring States period (B.C. 475- B.C. 221), Tang dynasty (A.D. 618- A.D. 907), Southern Song period (A.D. 1127- A.D. 1279), Ming dynasty (A.D. 1368- A.D.1644), and Qing dynasty (A.D. 1636- A.D. 1912) as well as some artificially aged silk specimens have been examined by scanning electron microscopy (SEM), attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy, and isotope ratio mass spectrometry (IRMS) to investigate their degradation for traceability and preservation. It has been found that the surface morphology and secondary structure of silk were destroyed in the degradation process. Under most aging modes, the silk fabrics were enriched with heavy stable isotopes after degradation. In the case of charring, the isotopic ratios of heavy hydrogen and oxygen isotopes showed different variation tendencies, which need to be further studied for better understanding. The results revealed that stable isotope ratios of silk fabrics are affected by different aging conditions. The degradation of silk fabrics is an important cause of stable isotope offset in historical silk fabrics. Overall, this study provides useful insights into the silk degradation mechanism and can serve as a useful reference for tracing the rich textile heritage. [ABSTRACT FROM AUTHOR]

[Display omitted] • Ultrasensitive detection is critical for investigating silk's origin and spread. • A dual-responsive PEC immunosensor was prepared for silk microtrace detection. • The sensor was developed by in-situ growth of Ag 2 S on ZnO-MXene nanocomposites. • The sensor exhibits excellent taxonomic specificity, repeatability and stability. • The dual-responsive PEC immunosensor is applied for detection of real silk samples. Developing an effective analytical method for microtrace detection of ancient silk is highly desirable and vitally crucial for exploring the origin and spread of silk. Herein, a taxonomically specific anti-fibroin monoclonal antibody was prepared and an elaborate visible/near-infrared (vis/NIR) photoelectrochemical (PEC) immunosensor was proposed. The sensor employed Ag 2 S nanoparticles (NPs) grown in situ on ZnO nanowire array-MXene nanocomposites (ZnO-MX) via a sonochemical method as a photosensitive material. The experimental results illustrated that the ZnO-MX/Ag 2 S-based sensor presented 6.8 times larger PEC responses than a ZnO-MX-based sensor under visible light illumination, and the PEC signal of the ZnO-MX/Ag 2 S nanocomposites was 30 times as high as that of ZnO-MX under 808 nm NIR light, indicating enhanced photoelectric activity and a matched energy band structure of ZnO with MXene and Ag 2 S. This PEC immunosensor exhibited excellent potential for the ultrasensitive detection of Bombyx mori silk traces due to its wide linear range of 0.005–50 ng mL−1 and low detection limit of 1.51 pg mL−1. Additionally, the immunosensor showed outstanding specificity, stability, and repeatability, particularly for the qualitative detection of archeological artifacts. Given its aforementioned benefits, ZnO-MX/Ag 2 S nanocomposites are potentially attractive photoactive materials for use in vis/NIR PEC immunosensors. Moreover, this demonstrated method offers a feasible method to investigate the history and distribution of ancient silk. [ABSTRACT FROM AUTHOR]

Ancient silk has important historical, artistic, and scientific value, and its origin and spread have always been a research hotspot. However, traditional analytical methods have encountered great challenges in the analysis of ancient silk microtraces due to the problems of large sample consumption, serious contamination, and degradation. To overcome this problem, a taxonomically specific anti-fibroin monoclonal antibody was designed and successfully prepared by animal immunization, and a novel photoelectrochemical immunosensor based on the antibody and ZnO nanowires array was constructed for the first time. This photoelectrochemical immunosensor possessed a detection limit as low as 0.56 ng mL−1 and a linear range from 1 to 1000 ng mL−1, showing great potential in the ultrasensitive detection of silk trace. In addition, the elaborate immunosensor demonstrated preeminent performances in the specificity, stability, and reproducibility, especially in the qualitative and quantitative detection of soil samples extracted from Sanxingdui archeological sites. Therefore, the proposed method not only provides a powerful analytical tool for exploring the origin and spread of ancient silk but also provides a reference for the study of other types of protein artifacts. • Ultrasensitive detection for silk microtraces has great significance to investigate the origin and spread of ancient silk. • Application innovation: it was the first time to design and prepare a PEC immunosensor in the archaeological field. • Method innovation: this work fabricated a PEC immunosensor based on CdS QDs and Ag NPs co-sensitized ZnO NWA. • The PEC immunosensor exhibited a low LOD of 0.56 ng mL−1 and was excellent in taxonomic specificity and stability. [ABSTRACT FROM AUTHOR]