Crystallography for biology - as easy as EBSD

Living systems exert exquisite control on the formation of their mineral structures producing structures that perform various functions such as protection in bivalve shells, embryonic chamber in bird eggs and skeletal support in vertebrates for example. To a great extent, the biological control exerted is poorly understood although the knowledge is highly sought after. In materials science and metallurgy, scientists probe complex materials in great details using a technique called Electron Back Scattered Diffraction (EBSD). EBSD involves firing a beam of electrons at a tilted sample. The planes within the structure of the samples, split the electron beam and the fragmented beam is bounced back and imaged on a screen. The image tells us what the structure is made of and its orientation. The applicant has been applying EBSD to the analysis of minerals produced by living systems to great effect. Already important information on the biomechanics of invertebrates has been obtained. In addition, clarification has been obtained regarding which questions should be addressed in order to improve our understanding of this biological control. The need for this application is apparent by the fact that many people are beginning to become aware of the huge potential of EBSD analysis to address questions in biological sciences. The need for this project is further heightened by the discovery by the applicant that EBSD analyses are not equally successful in all biominerals assessed thus far. This project aims to assess the important sample preparation conditions; resin type, duration of final polishing stage and thickness of carbon coat, in the shell of te hcommon blue mussel, Mytilus edulis which contains two polymorphs of calcium carbonate: calcite and aragonite. This initial survey of conditions will be used to define parameters in a wider biomineral survey that will include other calcium carbonate biominerals, brachiopods, corals and bryozoans as well as calcium phosphate (apatite) biominerals, namely the shell of the brachiopod Lingula anatina and the femur of commercially available domestic fowl. This larger survey will also quantify the organic and trace element composition so that the factors that may influence crystallography and its control may be identified in a range of biomineral systems. These two surveys will determine the optimal conditions for EBSD analysis of biogenic structures and will identify the limits of EBSD analysis. Both sets of knowledge are essential for the effective transfer of this powerful technique to biological sciences.

The proposal aims to assess the application of Electron Back Scatter Diffraction (EBSD) to a range of biominerals in order to gauge the breadth of the applicability of this technique to biogenic materials. The two major advantages of EBSD analysis over conventional techniques, such as X-ray diffraction of isolated components, is that EBSD provides highly detailed information down to a spatial resolution of 20nm and the data is obtained in context. However, preliminary analyses indicate that some biominerals are more difficult to analyse using EBSD than others. The reasons for this must be determined and the conditions optimised so that the advantages of EBSD analysis can then enable the chemistry and material properties to be understood in the context of crystallography providing detailed knowledge of biological control on structural components. The best possible sample preparation conditions for EBSD of biominerals will be determined by assessing preparation conditions using the shells of the common blue mussel, Mytilus edulis which contain two polymorphs of calcium carbonate; calcite and aragonite. Having established the optimal conditions for EBSD analysis, the technique will be applied to a range of carbonate and some apatite minerals. EBSD analyses of the wider biomineral survey will be done using longitudinal and transverse sections in order to determine the crystallography (EBSD), chemistry (EDAX and electron microprobe analyses) and organic content (loss on ignition and electron back scatter analysis) in 3-D. Thus, the inter-relationship between all of these factors will be determined in 3-D within the context of biomineralisation. This will greatly improve our understanding of biological control on mineral production in the context of detailed crystallography, determining the influence of organic and trace element composition on this control. This will assess the suitability of EBSD for applications in biological sciences.