Mammal Classification by Mass-Spectrometry Figure 2. MALDI-TOF MS peptide spectrum obtained from 8,500-year-old human dental pulp. doi:10.1371/journal.pone.0017319.g002 this study, we observed that the peptide spectra that were derived from the dental pulp contained several trypsin-digested proteins in addition to the collagen type I and collagen type III previously shown to be major constituents of the dental pulp [23]. We also found collagen type V - derived peaks, a collagen molecule also previously characterized in the dental pulp [23]. Peptide sequencing confirmed the detection of collagen in both modern and ancient dental pulp specimens, indicating that the dental pulp is a previously uninvestigated source for this protein which is widely used for the classification of ancient mammal individuals at the species level, in addition to bone tissue [7,24–27]. Moreover, peptide sequencing still proved the detection of 2 additional proteins in ancient dental pulp specimens, which have been previously reported as present in the human dental pulp [23,28,29]. Sequencing therefore indicated that peptide profiling identification did not rely on only one protein and that the dental pulp is a suitable source of non-collagen proteins. In our hands, accurate identification was obtained within 72 hours. These results indicate that MALDI-TOF MS peptide profiling could be used for rapid classification at the species level of mammalian remains collected from excavations, thus valuably contributing to the study of past human and non-human populations. Conclusions We observed that dental pulp samples were sufficient for both PCR-based sequence classification and MALDI-TOF MS peptide profiling in ancient and modern mammal individuals at the species level. Dental pulp is relatively easy to collect and is durable due to being embedded in solid teeth. Dental pulp is now potentially a new source for studying of ancient proteins, including collagen. Therefore, we recommend MALDI-TOF MS analysis of dental pulp as a reproducible approach complementing DNA sequencing for the accurate classification of forensic and ancient mammalian remains. Materials and Methods Sources of dental pulp We analyzed 13 modern mammalian species (37 total individuals, 37 teeth): cow (Bos taurus), goat (Capra hircus), camel (Camelus dromedarius), wild boar (Sus scrofa), pig (Sus scrofa subsp. domesticus), roe deer (Capreolus capreolus), rabbit (Oryctolagus cuniculus), rat (Rattus rattus), guinea pig (Cavia porcellus), cat (Felis catus), dog (Canis familiaris), red fox (Vulpes vulpes) and human (Homo sapiens). Human teeth were acquired from dentists after obtaining informed PLoS ONE | www.plosone.org 4 February 2011 | Volume 6 | Issue 2 | e17319