Focused ion beam milling and ultramicrotomy of mineralised ivory dentine for analytical transmission electron microscopy

The use of focused ion beam (FIB) milling for preparation of sections of mineralised ivory dentine for transmission electron microscopy (TEM) is investigated. Ivory dentine is essentially composed of fibrillar type-I collagen and apatite crystals. The aim of this project is to gain a clearer understanding of the relationship between the organic and inorganic components of ivory dentine using analytical TEM, in order to utilise these analytical techniques in the context of common skeletal diseases such as osteoporosis and arthritis. TEM sections were prepared in both single and dual beam FIB instruments, using two standard lift-out techniques, in situ and ex situ. The FIB sections were systematically compared with sections prepared by ultra microtomy, the traditional preparation route in biological systems, in terms of structural and chemical differences. A Clear advantage of FIB milling over Ultramicrotomy is that dehydration, embedding and section flotation call be eliminated, so that partial mineral loss clue to dissolution is avoided. The characteristic banding of collagen fibrils was clearly seen in FIB milled sections without the need for any chemical staining, as is commonly employed ill Ultramicrotomy. The FIB milling technique was able to produce high-quality TEM sections of ivory dentine, which are Suitable for further investigation using electron energy-loss spectroscopy (EELS) and energy-filtering TEM (EFTEM) to probe the collagen/apatite interface. (C) 2008 Elsevier Ltd. All rights reserved.