This study closely examines Domenico Cresti's oil-based wall paintings in Santa Maria della Pace in Rome using a novel analytical technique that allows for the simultaneous characterization of organic and inorganic materials in works of art. ToF-SIMS was successfully used to obtain high-resolution spatial maps of cross-sectional paint samples and served as a complimentary technique to Raman spectroscopy and scanning electron microscopy-energy dispersive X-ray spectroscopy. Analysis of Cresti's wall paintings using imaging ToF-SIMS revealed the presence of specific amino, acids as well as fatty acids, fragments that were mapped to discrete paint and ground layers. ToF-SIMS was also used to characterize proteinaceous materials that were presumably used during a previous restoration campaign. These analytical results allowed for direct comparison between Cresti's wall painting technique to those described primary sources, specifically those cited by Giorgio Vasari and Filippo Baldinucci.