Some 26 samples from the paint layer of the Last Supper have been analyzed by cross sections, microscopy, microchemistry, emission-spectrography, x-ray powder diffraction, x-ray fluorescence analysis induced by a proton source, and the scanning electron microscope. The plaster consists of two layers of lime mortar containing proteins and salts of fatty acids. The ground contains calcium carbonate and a hydrated magnesium carbonate bound by animal glue and small amounts of walnut oil. For the paint layer, soluble and insoluble proteins are the main constituents of the medium. Calcium-magnesium-carbonate, lead white, red and yellow iron oxide pigments, vermilion, minium, madder-lake, lead-tin-yellow, orpiment, green earth verdigris, azurite, indigo or woad, charcoal black, bone black, and soot have been identified as pigments. Gold, silver, and tin have been applied as leafs.