Skip to main content Accessibility help

Identification of Organic Binders in Ancient Chinese Paintings by Immunological Techniques

  • Wenjing Hu (a1), Hui Zhang (a2) and Bingjian Zhang (a1) (a2)


The identification and localization of organic binders in artworks are big challenges in archaeology and conservation science. Immunological techniques, such as enzyme-linked immunosorbent assay (ELISA) and immunofluorescence microscopy (IFM) have the potential to become powerful tools for the analysis of organic materials in ancient samples. In this study, ELISA and IFM techniques were combined to identify chicken ovalbumin, glue from several mammalian species, bovine milk, and fish glue in ancient Chinese painting samples. As binders, egg ovalbumin was found in two painting samples and animal glue was found in three samples, which were dated from the 4th to 8th centuries. The results clearly demonstrate that ELISA and IFM can be used to validate results from ancient samples.


Corresponding author

* Corresponding author.


Hide All
Arslanoglu, J., Zaleski, S. & Loike, J. (2011). An improved method of protein localization in artworks through SERS nanotag-complexed antibodies. Anal Bioanal Chem 399, 29973010.
Bonaduce, I., Blaensdorf, C., Dietemann, P. & Colombini, M.P. (2008). The binding media of the polychromy of Qin Shihuang’s Terracotta army. J Cultur Herit 9, 103108.
Cartechini, L., Vagnini, M., Palmieri, M., Pitzurra, L., Mello, T., Mazurek, J. & Chiari, G. (2010). Immunodetection of proteins in ancient paint media. Acc Chem Res 43, 867876.
Casoli, A., Musini, P.C. & Palla, G. (1996). Gas chromatographic-mass spectrometric to the problem of characterizing binding media in paintings. J Chromatogr A 731, 237246.
Colombini, M.P. & Modugno, F. (2004). Characterisation of proteinaceous binders in artistic paintings by chromatographic techniques. J Sep Sci 27, 147160.
Gambino, M., Cappitelli, F., Cattò, C., Carpen, A., Principi, P., Ghezzi, L., Bonaduce, I., Galano, E., Pucci, P., Birolo, L., Villa, F. & Forlani, F. (2013). A simple and reliable methodology to detect egg white in art samples. J Biosci 38, 397408.
Heginbotham, A., Millay, V. & Quick, M. (2004). The use of immunofluorescence microscopy (IFM) and enzyme-linked immunosorbent assay (ELISA) as complementary techniques for protein identification in artists’ materials. J Am Inst Conserv 45, 89106.
Hu, W., Zhang, K., Zhang, H., Zhang, B. & Rong, B. (2015). Analysis of polychromy binder on Qin Shihuang’s Terracotta Warriors by immunofluorescence microscopy. J Cultur Herit 16, 244248.
Kockaert, L., Gausset, P. & Dubi-Rucquoy, M. (1989). Detection of ovalbumin in paint media by immunofluorescence. Stud Conserv 34, 183188.
Kuckova, S., Sandu, I.C.A., Crhova, M., Hynek, R., Fogas, I., Muralha, V.S. & Sandu, A.V. (2013). Complementary cross-section based protocol of investigation of polychrome samples of a 16th century Moravian Sculpture by optical, vibrational and mass spectrometric techniques. Microchem J 110, 538544.
Kuckova, S., Sandu, I.C.A., Crhova, M., Hynek, R., Fogas, I. & Schafer, S. (2013). Protein identification and localization using mass spectrometry and staining tests in cross-sections of polychrome samples. J Cultur Herit 14, 3137.
Li, S. (1990). The application of HPLC technology on the analysis of mural binders. Dunhuang Res 4, 53.
Li, Z.X. (1990). The primary exploration of binders in Dunhuang murals. Dunhuang Res 3, 69.
Łojewski, T., Bagniuk, J., Kołodziej, A. & Łojewska, J. (2011). Reflective and photoacoustic infrared spectroscopic techniques in assessment of binding media in paintings. Appl Phys A 105, 753761.
Magrini, D., Bracci, S. & Sandu, I.C.A. (2013). Fluorescence of organic binders in painting cross-sections. Procedia Chem 8, 194201.
Mazzeo, R., Prati, S., Quaranta, M., Joseph, E., Kendix, E. & Galeotti, M. (2008). Attenuated total reflection micro FTIR characterisation of pigment-binder interaction in reconstructed paint films. Anal Bioanal Chem 392, 6576.
Palmieri, M., Vagnini, M., Pitzurra, L., Brunetti, B.G. & Cartechini, L. (2013). Identification of animal glue and hen-egg yolk in paintings by use of enzyme-linked immunosorbent assay (ELISA). Anal Bioanal Chem 405, 63656371.
Palmieri, M., Vagnini, M., Pitzurra, L., Rocchi, P., Brunetti, B.G., Sgamellotti, A. & Cartechini, L. (2011). Development of an analytical protocol for a fast, sensitive and specific protein recognition in paintings by enzyme-linked immunosorbent assay (ELISA). Anal Bioanal Chem 399, 30113023.
Potenza, M., Sabatino, G., Giambi, F., Rosi, L., Papini, A.M. & Dei, L. (2013). Analysis of egg-based model wall paintings by use of an innovative combined dot-ELISA and UPLC-based approach. Anal Bioanal Chem 405, 691701.
Richter, W., Krause, I., Graf, C., Sperrer, I., Schwarzer, C. & Klostermeyer, H. (1997). An indirect competitive ELISA for the detection of cows’ milk and caseinate in goats’ and ewes’ milk and cheese using polyclonal antibodies against bovine γ-caseins. Zeitschrift für Lebensmitteluntersuchung und -Forschung A 204, 2126.
Rosi, F., Federici, A., Brunetti, B.G., Sgamellotti, A., Clementi, S. & Miliani, C. (2011). Multivariate chemical mapping of pigments and binders in easel painting cross-sections by micro IR reflection spectroscopy. Anal Bioanal Chem 399, 31333145.
Sandu, I.C.A., Schäfer, S., Magrini, D., Bracci, S. & Roque, C.A. (2012). Cross-section and staining-based techniques for investigation of organic materials in polychrome works of art: A review. Microsc Microanal 18, 860875.
Schulz, H. & Kropp, B. (1993). Micro spectroscopy FTIR reflectance examination of paint binders on ground chalk. Fresenius’ J Anal Chem 346, 114122.
Spring, M., Ricci, C., Peggie, D.A. & Kazarian, S.G. (2008). ATR-FTIR imaging for the analysis of organic materials in paint cross sections: Case studies on paint samples from the National Gallery, London. Anal Bioanal Chem 392, 3745.
Su, B.M., Zhen, B.Z.F., Hu, Z.D. & Li, Z.X. (2005). HPLC analysis of binding medium of pigments of wall-painting at Kizil Grottoes. Dunhuang Res 4, 57.
Tokarski, C., Crenolive, C. & Rolando, C. (2003). Protein studies in cultural heritage, In Molecular biology and cultural heritage: Swets & Zeitlinger, Saiz-Jimenez, C. (Ed.), pp. 119130.
Vagnini, M., Pitzurra, L., Cartechini, L., Miliani, C., Brunetti, B.G. & Sgamellotti, A. (2008). Identification of proteins in painting cross-sections by immunofluorescence microscopy. Anal Bioanal Chem 392, 5764.
Wang, L., Yang, L., Zhou, W. & Fan, X. (2011). Comparison and evaluation of methods for quantitative analysis of proteins in wall paintings and colored relics of ancient architecture. Sci Conserv Archaeol 23, 5963.
Wei, S.Y., Ma, Q.L. & Schreiner, M. (2012). Scientific investigation of the paint and adhesive materials used in the western Han dynasty polychromy terracotta army, Qingzhou, China. J Archaeol Sci 39, 16281633.



Full text views

Total number of HTML views: 0
Total number of PDF views: 0 *
Loading metrics...

Abstract views

Total abstract views: 0 *
Loading metrics...

* Views captured on Cambridge Core between <date>. This data will be updated every 24 hours.

Usage data cannot currently be displayed