Nanoanalytical Electron Microscopy Reveals a Sequential

Nanoanalytical Electron Microscopy Reveals a Sequential Mineralization Process Involving Carbonate-Containing Amorphous Precursors Kharissa Nitiputri,†,‡,§ Quentin M. Ramasse,∥ Hélène Autefage,†,‡,§ Catriona M. McGilvery,† Suwimon Boonrungsiman,†,‡,§ Nicholas D. Evans,⊥ Molly M. Stevens,*,†,‡,§ and Alexandra E. Porter*,† †

Department of Materials, ‡Department of Bioengineering, §Institute of Biomedical Engineering, Imperial College London, London SW7 2AZ, United Kingdom ∥ SuperSTEM Laboratory, SciTech Daresbury Campus, Daresbury, Halton WA4 4AD, United Kingdom ⊥ Department of Bioengineering and Institute for Life Sciences, University of Southampton, Southampton SO17 1BJ, United Kingdom S Supporting Information *

ABSTRACT: A direct observation and an in-depth characterization of the steps by which bone mineral nucleates and grows in the extracellular matrix during the earliest stages of maturation, using relevant biomineralization models as they grow into mature bone mineral, is an important research goal. To better understand the process of bone mineralization in the extracellular matrix, we used nanoanalytical electron microscopy techniques to examine an in vitro model of bone formation. This study demonstrates the presence of three dominant CaP structures in the mineralizing osteoblast cultures: pi*co Transition in The Near-Edge X-ray Absorption Fine Structure Spectra of Organic Molecules. J. Phys. Chem. B 2002, 106, 8531−8538. (34) Brandes, J. A.; Wirick, S.; Jacobsen, C. Carbon K-Edge Spectra of Carbonate Minerals. J. Synchrotron Radiat. 2010, 17, 676−682. (35) Metzler, R. A.; Abrecht, M.; Olabisi, R. M.; Ariosa, D.; Johnson, C. J.; Frazer, B. H.; Coppersmith, S. N.; Gilbert, P. U. P. A. Architecture of Columnar Nacre, and Implications for Its Formation Mechanism. Phys. Rev. Lett. 2007, 98, 268102. (36) Rey, C.; Kim, H. M.; Gerstenfeld, L.; Glimcher, M. J. Characterization of The Apatite Crystals of Bone and Their Maturation in Osteoblast Cell Culture: Comparison with Native Bone Crystals. Connect. Tissue Res. 1996, 35, 343−349. 6834

DOI: 10.1021/acsnano.6b02443 ACS Nano 2016, 10, 6826−6835

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ACS Nano (55) Garvie, L. A. J. Can Electron Energy-Loss Spectroscopy (EELS) be Used To Quantify Hydrogen in Minerals from the O K edge? Am. Mineral. 2010, 95, 92−97.

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DOI: 10.1021/acsnano.6b02443 ACS Nano 2016, 10, 6826−6835