With recognition of the early Holocene antiquity of marine shell beads in Island Southeast Asia only recently occurring, we become aware of how little is really known regarding this enigmatic class of material culture. Here we report on worked Nassarius spp. shells recovered from the Timorese sites of Jerimalai, Lene Hara, and Matja Kuru 1 and 2, and which date back to around 6500 years ago. Analysis of manufacturing traces, use wear, and residues apparent on these 91 shell artefacts indicate that they were most likely used as appliqués attached to a textile or other woven item (such as baskets). These are the first mid-Holocene shell appliqués to be identified in this region, and only the second example of this technology at this antiquity identified in the world. Consistency in manufacturing methods and use over several thousand years at the studied sites indicates a >4500 year long tradition of Nassarius spp. shell appliqué use in Timor-Leste.