Excellently preserved animal bone and antler artifacts are numerous among the archaeological finds in Neolithic Alpine lakeshore settlements. Use-wear and production traces on the surface of these artifacts include cuttings and hammer or grinding traces; there are also large bright and shiny areas. We have explored the origin of such shiny surfaces using molecular and isotopic analysis of the lipids staining freshly unearthed and non-conserved bone and antler artifacts from a recent excavation at the underground parking garage of Zurich Opera House (Switzerland), dating to ca. 3230–2729 BC. A set of 10 artifacts, covering distinct osteological and typological features, was selected for study. The lipids were extracted separately from the surfaces of the tapered thinner working part and the wider handling area of each tool. The lipid distribution was characterized by a significant amount of animal sterols (cholesterol, cholestanol, coprostanol and epicholestanol), wax n-alkanes, n-alkan-1-ols and phytosterols (β-sitosterol, sitostanol) and abundant saturated, mono- and polyunsaturated fatty acids (FAs) in the C14–C26 range. The δ13C values of the main FAs, and the relatively high C18:2 content (up to 31% of total FAs) and significant content of C18:3(up to 6%) indicated that the lipids on the surface of the artifacts were a mixture of C3 plant oils rich in linoleic acid and (indigenous) animal lipids. The results provide evidence for the archaeological hypothesis that the bone and antler artifacts were specific tools which were deliberately fashioned, and pretreated and maintained with a preservative material based on plant oil likely from seeds of Linum usitatissimum (flax) andPapaver somniferum (poppy), with probably some contribution from Corylus avellana (hazelnut) andBrassica rapa (turnip).