In order to clarify more completely the morphophysiology of hard tissues, it is of foremost importance to determine the precise location and the elements of the extracellular and cellular mineral constituents. In the present study we have avoided the potential artefacts which may be introduced by the routine preparation for electron microscopy by adapting both anhydrous and frozen ultrathin-sectioning techniques to our fine structural and elemental analytical studies on hard tissues.Freshly dissected developing epiphyseal cartilage, bone and dentin of tooth germs from young postnatal rats have been prepared for the analytical electron microscopy(Hitachi H500H Total System equipped with Kevex 7000 X-ray Energy Spectrometry) by the methods of the ultracryo- microtomy(Christensen,1971) and the inert dehydration technique(Pease, 1966.,Landis et al, 1977).Nuclei, ribosomes, endoplasmic reticulum and a large number of dense intramitochondrial granules, 20 to 80nm in diameter, within the chondrocytes(Fig. 1a,b), osteoblasts(Fig.2) and odontoblasts(Fig.3) were observed in unfixed, unstained frozen ultrathin-sections, obtained with dry knives in the temperature range of -60 to -80°C, and also in unstained sections treated anhydrously by ethylene glycol.
Spinel-type lithium titanate (Li 4 Ti 5 O 12 (LTO)) is known as a negative electrode material for lithium-ion batteries (LIBs). In this study, the surface structure of the LTO(111) electrode in an ionic liquid (IL) electrolyte was investigated by electrochemical atomic force microscopy (EC-AFM). A quartz tuning fork sensor with a sharpened tungsten tip, so-called the qPlus sensor, was used as an AFM force sensor instead of a Si cantilever, and the AFM equipment was embedded in a vacuum chamber to decrease the water concentration included in the IL electrolyte. The in situ analysis of the surface structure before and after the insertion of Li ions into the LTO electrode revealed that the atomic steps on the LTO(111) surface were kept even after the insertion of Li ions. We also succeeded in atomic-resolution topographic imaging on the LTO(111) electrode before and after Li-ion insertion. The hexagonally arranged bright spots with a period of ∼0.3 nm were clearly imaged after Li-ion insertion, which corresponded well to the structure of the ion arrangement of the Li 7 Ti 5 O 12 (111) surface. Namely, after Li-ion insertion, it was revealed that the LTO electrode changed its crystal structure from spinel-type Li 4 Ti 5 O 12 to rocksalt-type Li 7 Ti 5 O 12 while maintaining the surface structure with atomic steps.
A two-step sharpening process for a Si pillar in a quartz tuning fork sensor was developed for use in frequency modulation atomic force microscopy (FM-AFM). By combining electrochemical etching in HF solution and anisotropic etching in KOH solution, the tip radius was decreased to 120 nm. The Si-probe-attached sensor showed a higher resonance frequency than a tungsten-probe attached sensor, which would lead to a higher force sensitivity. We demonstrated FM-AFM imaging on a cleaved mica surface in aqueous solution by using the fabricated sensor, and atomic resolution was successfully achieved.
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