Proteomic sexing of archaeological cattle remains at Neolithic Kilshane

Although proteomic techniques have been increasingly used to improve our understanding of the human past, few have focussed on the study of tooth enamel for sexing in archaeofaunal remains, despite initial studies over a decade ago investigating human teeth. Here we explore the use of LC-orbitrap-MS/MS for identifying the sex of archaeological domestic cattle remains from the Neolithic enclosure at Kilshane, Ireland (c. 3600 cal. BC), in addition to modern reference materials. Although several individuals could be confidently identified as male, and some less confidently as female, there were also some inconsistencies with predictions based on morphology, albeit such estimations based largely on size. Through comparison with osteometric analyses, the proteomic analyses estimated 14 consistent identifications and six discrepancies, with one of the two speculative males confirmed through the use of proteomics. The male/female proportions for the osteometric analysis based on the metacarpals, the most sexually dimorphic skeletal element, showed that slightly over 50% of the bones were from females and slightly under 50% from males, whereas in the case of proteomics, depending on what level of caution we place on biomarker usage, this is much lower for the identification of males, ranging from 12.5 to 46% (3–11 specimens of 24 successful proteome samples with estimated sex based on their molars). Unexpectedly, our results show evidence of clear differences between modern reference materials and archaeological materials, likely reflecting preservation biases relating to the protein sequences.