## 3 Results  
The cross-sections analyzed under the UV-light of the microscope show the presence of four distinct layers as indicated in Figure 2d: (1) an uppermost white-opalescent layer, 40 \(\upmu\)m thick approximately; (2) a thinner (circa 15 \(\upmu\)m) fluorescent orangish layer; (3) a very thin (submicron) black layer, likely ascribable to the silver leave; and (4) a very thick whitish preparation layer showing (Figure 2a,b) a double row of holes left by the canvas, which was applied throughout the panel and on the frame by means of a mixture of chalk and glue in order to cushion the movements of the wood. This procedure is in line with the preparatory technique of that time justifying also the considerable thickness of the ground. Analysis carried out on the canvas led to rough linen with a coarse \"plain weave\" texture (weft/warp ratio 1: 1). Layers (1) and (2) were distinctly observed only with the UV-light. The same layers observed under VIS-light Optical Microscopy (OM) appeared as a unique layer of yellow color (see Figure 3a). This suggests that its function was likely just to make silver brighter and probably like gold, as clearly displayed in Figure 3b. Noteworthy is the \"wavy\" form of the orangish layer (layer 2), which is likely ascribable to the grooves left by bristles of brush used to apply layer 1 (see Figure 2c,d). A better knowledge of the chemical composition of the different layers might provide crucial information to shed light on these issues. The analysis of cross-sections by UVIVF micro-spectroscopy revealed to be very supportive for the discrimination of the two kinds of varnish layers found on top of the silvered surface. Normalized UVIVF spectra collected on the organic layers over the silver leaf are shown in Figure 4.  
Figure 3: VIS-light image of a painting sample as seen under Optical Microscopy (OM): **(a)** cross-section of the gilded surface; **(b)** surface with the varnished (yellow) and unvarnished (gray) silver (Ag) layer. Figure 2: UV fluorescence microscope images of a painting cross-section at different magnification objectives: **(a)** 10\(\times\), **(b)** 20\(\times\), and **(c,d)** 100\(\times\). Four different layers were observed: (1) a white-opalescent layer (circa 40 \(\mu\)m); (2) an orangish layer ( circa 15 \(\mu\)m); (3) a sub-micron black layer; and (4) a very thick whitish preparation layer. Fluorescence maxima (\(\lambda_{\text{max}}\)) were found at about 500 (136 nm, measured at Full Width Half Maximum (FWHM) of the band) and 580 nm (230 nm FWHM) for the varnish layer 1 and 2, respectively, according to the scheme shown in Figure 2d. Fluorescence intensity was strongly lower for layer 2 applied over the silver leaf in comparison to the thicker varnish on top (i.e., layer 1), thus suggesting that layer 2 was more aged and ancient. As witnessed by the narrower FWHM, varnish layer 1 is instead more representative of a less complex and aged composition [13]. Unlike varnish layer 2, the blue-shifted fluorescence of the uppermost varnish layer 1 could match more plausibly with a yellow oil medium glaze, but it is rather difficult to obtain more detailed information. From qualitative comparisons of DRIFT and \(\upmu\)-ATR FT-IR spectra additional and valuable information on the artistic technique were pointed out. In detail, DRIFT analysis of paint fragments before being embedded is shown in Figure 5A. Starting from the top (Figure 5A, spectrum a), the broad C=O stretching band in the 1745-1730 cm\({}^{-1}\) region with a shoulder at 1778 cm\({}^{-1}\) and the C-H bending at 1465 cm\({}^{-1}\) were indicative of an oil-based glaze. Signals near 2900 cm\({}^{-1}\) (C-H stretching band) were scarcely informative, whereas the clear shoulder-peak at 1676 cm\({}^{-1}\) (C=O stretching band) could be from amide I in proteins. The evidence of the latter was found also in other fragments extracted from the same sampling point (Figure 5A, spectrum b).