The late nineteenth and early twentieth centuries were marked by tremendous innovations in art and science. On the one hand, with the rapid rise of the chemical industry, numerous new inorganic and organic pigments were developed and marketed as alternatives to wellestablished traditional pigments. On the other hand, development of spectrophotometry and the flourishing of numerous treatises on color theory, were soon made available to artists. Equipped with a new prismatic palette and new scientific theories, Neo-Impressionist artists extensively discussed and exploited color mixtures, and by using a variety of painting techniques, they created new facture. In this context, the in-situ analytical study of Paul Signac painting materials and techniques represents a unique opportunity to determine the relationships between nineteenth century literature on color theory and its application by the contemporaneous Neo-Impressionist painter.