The two layers of the shell of a living fossil, Neotrigonia, were examined using optical, electron and atomic-force microscopies. Nacre and prisms are composed of complex acicular crystallites. These crystallites are heterogeneous and composed of rounded granules surrounded by a thin cortex. Despite both layers being aragonite, the chemical compositions of the layers differ as shown by in situ electron microprobe analysis. XANES maps show that sulphur is linked to the organic components. The external part of the prismatic layer is rich in sulphur aminoacids and organic sulphate, as well as in Mg, Sr and P. The outer part of the nacreous layer is enriched in sulphur aminoacids. Such differences have been described in aragonitic–calcitic shells, but not in a single-mineral shell. This nacreous arrangement is not typical of Bivalve shells.