Though ancient violins and other stringed instruments are often revered for the beauty of their varnishes, the varnishing techniques are not so much known. In particular, very few detailed varnish analyses have been published so far. Since 2002, a research program at the Musée de la musique (Paris) is dedicated to a detailed description of varnishes on famous ancient musical instruments using a series of novel analytical methods. For the first time, results are presented on the study of the varnish from a late 16th century Venetian lute, using synchrotron micro-analytical methods. Identification of both organic and inorganic compounds distributed within the individual layers of a varnish microsample has been performed using spatiallyresolved synchrotron FTIR microscopy. The univocal identification of the mineral phases is obtained through synchrotron powder X-ray diffraction. The materials identified may be of outmost importance to understand the varnishing process and its similarities with some painting techniques. In particular, the proteinaceous binding medium and the calcium sulfate components (bassanite and anhydrite) that have been identified in the lower layers of the varnish microsample could be related, to a certain extent, to the grounds materials of earlier Italian paintings.