# High Resolution STEM Images of the Human Tooth Enamel Crystals
## 1 Introduction
### The Central Dark Line  
At the central region of the human tooth enamel crystals, a defect named as \"the central dark line\" (CDL) was observed. In transmission electron microscopy (TEM) images, the CDL showed a contrast similar to the \"Fresnel contrast\" of edge-on defects; that is, it appeared as a dark line with a width of ~1 nm out-of-focus in both cross-sectional and transverse section samples, but it was white in over-focus and disappeared in-focus (the Gaussian focus) [3; 4; 5]. Figure 1 shows a bright-field TEM image of human tooth enamel crystals, where the arrows indicate the positions of the CDL. The inset shows a high-resolution TEM (HRTEM) image of the CDL in one of these crystals. Because enamel crystals begin to dissolve from the center during acidic attacks such as caries [6], the CDL is of particular interest. Figure 2a shows the graphic representation of the dissolution of the human tooth enamel by acid attack, and Figure 2b shows its HRTEM image as they began to dissolve after treated with orthophosphoric acid for simulating acid attacks: the acid attack always started at the center of the crystals. Taking as a basis the hexagonal unit cell of HAP, during caries, the human tooth enamel crystals were destroyed systematically: first, a central lesion elongated along the [11; 12; 13; 14; 15; 16; 17; 18; 19; 20] direction appeared on the basal (0001) planes; and secondly, the lesion developed anisotropically along the [0001] direction across the crystals [7]. The TEM observations of the enamel crystals indicated that the CDL was in fact a planar defect. Several hypotheses have been proposed in its possible origin. One of them is that the CDL is a transformation-mismatched layer from octacalcium phosphate (OCP) to HAP. However, the HAP-OCP interface model generated does not fully reproduce the TEM data [8]. Another possibility is that the CDL is a Ca-rich region within enamel crystals and therefore cannot be the residual of the OCP-HAP mismatch [8; 9]. The contrast of the CDL, when observed with a scanning transmission electron microscope (STEM) using an annular detector, gave additional clues on the CDL's structure. Modern STEM microscopes were equipped with aberration correction systems, and the low- and high-angle scattered electron annular detectors provided bright-field (BF), low-angle annular dark-field (ADF), and high-angle annular dark-field (HAADF) images with sub  
Figure 1: TEM image of human tooth enamel. The Inset shows the high-resolution TEM (HRTEM) image of one of these crystals. The arrows indicate the central dark line (CDL) positions. angstrom resolution [10]. STEM-BF images include the transmitted beam and small-angle scattered beams produced by Rutherford elastic scattering, plasmons, and phonons [11]. When human tooth enamel crystals were observed by an STEM, the BF image showed a contrast similar to the HRTEM images, and the CDL appeared dark (Figure 3a). In the STEM-ADF image, the collection of scattered beams was made at angles between 25 and 250 mrad and, therefore, their contrast present elastic deformation fields (Figure 3b) [11]. In this case, the CDL showed a bright contrast surrounded by an extended gray zones produced by strain (Figure 3d) [2,3]. For the HAADF-STEM image, scattered beams at angles between 50 and 250 mrad were collected. These large-angle scattered beams were related to the atomic number of the sample (the Z-contrast), and the contrast of these images at atomic resolution was \"interpretable\" in a direct way [11]. That is, the light contrast indicated atomic positions with a larger Z value, and the gray and dark contrast were related to atomic positions with a smaller Z value. Figure 3c shows the HAADF-STEM image of the human tooth enamel crystal at a low magnification. In this case, the CDL contrast was weak and, sometimes, becomes almost invisible, which indicated that the large-angle scattered beams were generated in practically the whole crystal.