J. Anat. (1978), 125, 2, pp. 323-335 With 27 figures Printed in Great Britain
The enamel-dentine junction of human and Macaca irus teeth: a light and electron microscopic study D. K. WHITTAKER
Department of Oral Biology, Welsh National School of Medicine, Dental School, Heath, Cardiff
(Accepted 20 January 1977) INTRODUCTION
Junctional zones, particularly those between tissues of differing physical properties, present problems to the histologist and this may explain the rather sparse literature on the structure of the enamel-dentine junction. An early description of the junction was given by Tomes (1898), who implied that all enamel is festooned towards the dentine surface. Few workers have questioned this concept although Rywkind (1931) noted that in some teeth the scallops are irregular in size and distribution and are sometimes absent. Gustafson (1961) confirmed the basic arcade-shaped appearance of the junction, but agreed that the development of the scallops varies from tooth to tooth, and suggested that the pattern may be characteristic of the individual. She noted that the arcades were more pronounced in fluorosed teeth. Falin (1961) described the junction in Bronze Age teeth as being flat or slightly festooned in premolars and molars, and scalloped in canines and incisors. No comparable study appears to have been carried out in teeth of modern origin. More recently Scott & Symons (1974) commented upon the variation in size of the dome-shaped scallops, which are usually most marked in the cuspal region, but are occasionally absent. Their views are at variance with those of Schour (1960), who claimed that scalloping is more marked in the gingival third of teeth. No general agreement exists as to the size of the scallops (Nylen & Scott, 1958; Masukawa, 1959; Nakamura, 1959), and there has also been debate concerning the relationship of the crystals of enamel and dentine at the junction (Helmcke, 1953; Heuser, 1961; Nalbandian & Frank, 1962; Takuma, 1967; Watson & Avery, 1954). A further controversy has concerned the details of the development of the junction, which was originally thought to be produced by resorption of the dentine surface (Walkhoff, 1924). The central issue of the current debate is whether or not scallops are produced during the soft tissue stage of tooth formation (Schour, 1960) or only at a later stage (Provenza, 1964). The present study on both human and animal teeth seeks to resolve some of these problems. MATERIALS AND METHODS
The enamel-dentine junction (EDJ) was studied in non-carious teeth from both deciduous and permanent human dentitions and those of Macaca irus monkeys. The macaques were obtained as preserved post mortqm specimens from the Medical Research Council Laboratories, London. In addition human fetal teeth were examined. The specimens were fixed in 10 % formol saline and prepared for light microscopy using the following techniques: 2I-2
D. K. WHITTAKER
Table 1. Techniques and numbers of specimens studied Permanent
Preparation technique Specimens Human fetal Araldite embedded (a) Decalcified and sectioned Human non-carious (b) Ground (c) SEM of decalcified dentine (d) Fractured through ADJ (i) Embedded (ii) Not embedded (e) Ground, etched and SEM (f) Separated at ADJ and SEM (a) Decalcified and sectioned Monkey Macaca irus (b) Ground (c) SEM of decalcified dentine (d) Ground, etched and SEM Total
6 6 5
4 4 3
8 8 7
4 6 6 4
4 6 4 4
4 3 4 3 32
4 2 4 2 49
4 3 4 3 26
4 3 4 3 55-162
Anterior 9 3 3 3
(1) Teeth were decalcified in formic acid with continuous agitation until radiographic examination indicated total removal of inorganic material. The specimens were ultrasonicated to ensure removal of enamel matrix debris, dehydrated and embedded in paraffin wax. 5 ,um sections cut at right angles to the junction in both transverse and longitudinal planes were prepared and stained with either haematoxylin and eosin or Schmorl's picrothionin. (2) Undecalcified human fetal teeth were embedded in Araldite, sectioned at 1 4am and stained with toluidine blue. (3) Ground sections in longitudinal and transverse planes were prepared and mounted in Canada balsam. Specimens prepared by all these techniques were examined in a Leitz Wetzlar microscope with a x 100 objective and fitted with a camera lucida attachment. Drawings were made at standard magnification of representative zones of the junction from each specimen. Further specimens of human and monkey teeth were prepared for electron microscopy as follows: (1) Ground sections in longitudinal and transverse planes were prepared. The polished surfaces were etched with 0-25N HCl for 15 seconds and then washed and dehydrated. Positive replicas of the EDJ region were obtained using a two stage cellulose acetate and carbon technique (Bradley, 1957). Cellulose acetate strips were softened in acetone, pressed on to the etched tooth sections and allowed to dry. They were gently peeled off and vertically coated with carbon in an AEl Metrovac Type 12 vacuum coating unit. The acetate strips were dissolved by floating on acetone, and the carbon replica picked up from the surface of the solvent on copper grids. These replicas were examined in an AEl EM6B transmission electron microscope (TEM). The etched sections were mounted on aluminium stubs and coated by evaporation of gold in a Polaron E500 diode sputtering system at a vacuum of 0-07 Torr. Coating was continued for 2 minutes in an Argon gas atmosphere. Specimens were examined at various magnifications in an ISI Super Mini SEM. (2) Undecalcified teeth were embedded in Araldite, cut through from the dentine to within 1 mm of the EDJ, and then fractured through the junction. The fractured
Structure of the enamel-dentine junction 325 surfaces were ultrasonicated to remove debris, coated with gold and examined in the SEM. (Mortimer & Tranter, 1971). (3) The dentine surfaces of decalcified teeth were gold coated and examined in the SEM. (4) Enamel and dentine were separated at the EDJ by preparing 1 mm sections at right angles to the junction and desiccating them until splitting occurred. Both the exposed enamel and dentine surfaces were gold coated and examined in the SEM. Enamel was also separated from dentine by prolonged immersion of 1 mm tooth sections in sodium hypochlorite until dentine could be carefully dissected away from the enamel. Enamel surfaces exposed in this way were examined in the SEM. The number of teeth examined by these techniques is shown in Table 1. RESULTS
Contour of the junction in developing teeth In the monkey material the junction showed concavities which were approximately 5 ,am in diameter, and each was related to a prism end (Fig. 1). Apart from these small concavities the junction was not scalloped. The junction in human teeth varied in appearance with the stage of development. Before hard tissue formation some evidence of scalloping was visible at the basement membrane adjacent to the internal enamel epithelium (Fig. 2). Shortly after dentine and enamel formation began but before maturity was reached, evidence of scallops was rare and the junction usually appeared flat (Fig. 3). Contour of the junction in mature teeth In the human material the contour varied with the particular tooth examined, and also within the same tooth (Fig. 4). Scalloping was present in all the teeth in some sites (Fig. 5), but the amplitude and depth varied, and scalloping was absent or markedly reduced near the enamel-cement junction of most teeth (Fig. 6). Small scallops were seen in the permanent premolars and molars, deciduous molars and anterior teeth. The largest scallops were seen in the permanent anteriors. The depth of the scallops appeared to be less in the deciduous than in the permanent teeth, and the pattern of scalloping varied according to the tooth examined. Permanent and deciduous molars had the largest size of scallops in the cuspal region. In deciduous anteriors the distribution of the scallops appeared to be random, whilst in the permanent anteriors the largest scallops were seen in the cingulum and approximal portions of the teeth (Fig. 7). In the monkey material true scalloping was rare, and usually found only in approximal surfaces of the teeth. Elsewhere the junction was grossly flat, but undulated in 5 4tm concavities apparently related to the prism ends (Figs. 8 and 9). The larger true scallops were less well developed than in human teeth and appeared to be shallower. Surface features of the dentine SEM examination of the dentine surface in decalcified teeth enabled the morphology of the junction to be studied (Fig. 10). The variation in size of the scallops between teeth and within the same tooth was confirmed in the human material. The scallops consisted of crater-like depressions in the dentine surface (Fig. 11), the floor of which was composed of a dense network of fibrous-like material perforated by holes of varying size and having a roughened irregular texture. The
D. K. WHITTAKER
Fig. 1. Developing monkey permanent molar. Scallops at junction (arrowed) are 4-5 ,um in size and associated with enamel prisms seen in enamel matrix. H & E. x 300. Fig. 2. Developing human tooth before calcification. Note the slight scalloping of the future enamel-dentine junction (arrowed). H & E. x 300. Fig. 3. Developing human tooth after initial dentine (D) and enamel (E) formation. Note absence of scallops at junction (arrowed). H & E. x 300. Fig. 5. Ground section of a typical human permanent tooth. Note scallops at the junction. Ground section. x 300. Fig. 6. Human tooth showing junction near enamel-cement margin. Note absence of scallops. Ground sections. x 300. Fig. 7. Human permanent anterior tooth. Scallops at cingulum are large. Ground section. x 300. Fig. 8. Ground section of a typical tooth from Macaca irus. Large scallops are absent, but small concavities are present in dentine (arrowed) and relate to prism ends. Ground section. x 300. Fig. 9. Dentine from decalcified monkey tooth. Small scallops are visible (cf. Figs. 1 and 8). H&E. x 300.
Structure of the enamel-dentine junction A
Fig. 4. Camera lucida tracings of enamel-dentine junctions. Cross hatched structure represents dentine. 1, monkey molar; 2, human permanent molar; 3, human deciduous molar; 4, human anterior teeth. A, near cusp tip; B, near cervical margin; C, buccal surface, except C4 cingulum.
boundaries of the scallops consisted of raised ridges composed of more densely packed material (Fig. 12). The raised margins of adjacent scallops were fused, resulting in a continuous reticulum over the dentine surface. The dimensions and outline of the scallops were variable, ranging from circular to triangular or rectangular, and discrepancies in the scallop boundaries resulted in continuity in some places between adjacent scallops. Partial septae crossing the floor of the concavities were also present (Fig. 13). Towards the cervical margin of the teeth the raised boundaries of the scallops became more flattened, and the scallops elongated, resulting in flattened areas of dentine crossed by poorly developed ridges running longitudinally (Fig. 14). Differences in size, shape and distribution of scallops were clearly evident on adjacent surfaces of the same tooth. No clear differences were discernible between the human deciduous and permanent teeth, the size, shape and distribution of scallops being similar in both groups (cf. Figs. 15 and 13). In the monkey material scalloping was usually absent (Fig. 16). Scalloping appeared to be present only in the approximal surfaces of the teeth and was poorly developed compared with the human material (cf. Figs. 17 and 15). The shape and size of the depressions were similar to those in human teeth, but they were shallower, with thinner and narrower raised borders. In the base of each depression were hexagonal secondary depressions (Fig. 17). These were similar to those seen in nonscalloped areas, and were approximately 5 ,tm in size. No differences were noted between deciduous and permanent teeth in these respects.
D. K. WHITTAKER
Structure of the enamel-dentine junction
Fractured and ground sections through enamel-dentine junction These sections were prepared as nearly as possible at right angles to the junction in human deciduous and permanent teeth. Specimens which were not embedded in Araldite showed separation of the enamel and dentine at the junction. These were discarded, and only embedded material studied. It was not possible to distinguish between deciduous and permanent teeth so far as the structure of the EDJ was concerned. In human teeth the enamel prisms were closely applied to the dentine, but in some teeth of both permanent and deciduous series there was a zone of altered enamel about 20 ,um in width in contact with the junction (Fig. 18). In the acid-treated specimens this latter appeared to have been more heavily etched than the remaining enamel and the structure of individual prisms was less distinct. The zone of altered enamel was also visible on positive replicas of the junction in both deciduous and permanent human teeth and it appeared that the crystals within the prisms adjacent to the junction were fewei- and less well orientated than those in the remaining enamel (Fig. 19). In the fractured material the zone, when present, appeared as a homogeneous area where prism outlines were indistinct (Fig. 20). The relationship of dentinal tubules to the junction was most clearly seen in the fractured specimens. Some branching occurred near the junction, but not excessively so. The majority of the tubules terminated at or just before the junction, but occasional tubules appeared to enter the enamel. No evidence of enamel spindles was seen in specimens prepared in this manner but enamel tufts were obvious in the etched specimens and appeared as clefts between th. prisms (Fig. 21), occasionally extending across the junction into the dentine. Prisms associated with the tuft sides appeared to be disorientated. The junction between enamel and dentine at the cervical margin of the tooth was flattened and close interdigitation between the two tissues was apparent. Enamel in this zone was irregular in structure, some of the prisms appearing as flattened plates (Fig. 22). Etched ground sections through the junction of the monkey teeth showed a variable pattern. In those parts of the tooth where shallow scallops were present there was a zone of altered enamel about 12 ,um in width (Fig. 23 a) in which prism outline was indistinct. This appearance was usually found in approximal surfaces of the teeth. Elsewhere the scalloping was absent, but 5 ,um depressions associated with the convex prism ends were seen (Fig. 23b). Fig. 10. Low power scanning electron micrograph (SEM) of dentine surface of decalcified human tooth. Note variation in scallop dimensions. x 30. Fig. 11. Crater-shaped scallop on dentine surface of human permanent molar. SEM. x 1000. Fig. 12. Boundaries of scallops in a deciduous human tooth. Decalcified dentine surface. Note density of boundaries compared with floor. SEM. x 1000. Fig. 13. Pattern of scallops in a human deciduous molar tooth. Note incomplete crater walls and irregular shapes of scallops. SEM. x 200. Fig. 14. Human molar dentine surface close to enamel-cement junction. Note lack of scallops but presence of raised ridges. SEM. x 1000. Fig. 15. Human permanent molar tooth. Note similarity of scallops to those in deciduous teeth (Fig. 13). SEM. x 400. Fig. 16. Monkey permanent molar. Scallops are few in number, less concave, and with poorly marked boundaries. Depressions of enamel prism ends are visible. Decalcified dentine. SEM. x 400. Fig. 17. Monkey permanent molar. Most areas have no scallops, but imprint of enamel prism is visible. SEM. x 500.
D. K. WHITTAKER
Structure of the enamel-dentine junction
Surface features of the enamel Clean separation of enamel and dentine was achieved using the described techniques. In the human teeth convexities on the enamel surface were seen in areas of scalloping and the size of these convexities corresponded to the concavities in the dentine surface (Fig. 24). Fissures between them corresponded to the raised edges of the dentinal scallops. The ends of the prisms were visible on the enamel surface, but their outline appeared to be blurred by a surface coating. At fractured edges of the specimens the enamel prisms could be seen to be extending to the junctional surface. Occasionally tube-like structures 2 ,um in diameter were visible in the enamel surface. These were either situated within a prism (Fig. 25) or between prisms (Fig. 26). In the latter case their somewhat hexagonal walls were contributed to by numerous prisms. In the areas of the tooth where no scallops were present the ends of the prisms were concave (Fig. 27). DISCUSSION
In previous studies there has been disagreement concerning the details of the development of the enamel-dentine junction. Early workers (Walkhoff, 1924; Meyer, 1925) were of the opinion that, shortly after dentine formation had begun, the surface was resorbed resulting in depressions analogous to Howship's lacunae. These were then filled as enamel was deposited. Tylman (1928) and Orban (1929) clearly demonstrated the fallacy of this argument and Rykwind (1931) presented evidence, from studies of an odontome, that buds of the internal enamel epithelium result in a scalloped junction before calcification takes place. Controversy still exists as to the timing of scallop formation. In the monkey material the concavities in the dentine surface are related to the convex or-dorneshaped ends of the enamel prisms, but the fact that the mantle dentine is laid down prior to enamel formation (Reith & Butcher, 1967) suggests that sufficient plasticity remains in the newly formed dentine for the enamel to imprint its structure into the dentine as prisms are formed. This ability of the dentine surface to distort appears to be carried to even greater lengths in the human tooth since the present studies have shown that, although there is waviness of the basement membrane between internal enamel epithelium and odontoblasts, there is little evidence of Fig. 18. Human deciduous molar. Ground and etched section through junction. Note apparent fusion of enamel prisms close to junction. SEM. x 1000. Fig. 19. Ground, etched and replicated section through human permanent molar. Note sharp boundary between enamel (E) and dentine (D) and zone of altered enamel adjacent to junction. Transmission electron micrograph. x 800. Fig. 20. Human deciduous tooth fractured through junction. Note homogeneous nature of prisms close to junction, termination of dentinal tubules, and sharp boundary between enamel and dentine. SEM. x 500. Fig. 21. Ground, etched section of permanent human premolar. Note enamel tuft and its extension into dentine. SEM. x 400. Fig. 22. Enamel (E) and EDJ (arrowed) close to enamel cement junction. Human permanent molar. Note irregularity of enamel structure. SEM. x 400. Fig. 23. Ground and etched section of monkey tooth. (a) Approximal portion. Shallow scallops are visible with an altered zone of enamel in contact with the junction. SEM. x 550 (x 400). (b) Non-scalloped zone. Prisms commence at the junction and 5 ,um concavities in the dentine (arrow) relate to their ends. SEM. x 400.
D. K. WHITTAKER
Fig. 24. Human permanent molar. Junction surface of enamel. Convexities are comprised of numerous prism ends and sulci of varying depths separate them. Prisms are seen in longitudinal section in fractured surface (F). SEM. x 400. Fig. 25. Junction surface of enamel perforated by tubule for enamel spindle. Note that prism (P) was deposited around the spindle. SEM. x 4000. Fig. 26. Preparation as in Fig. 25, but note that several prisms contribute to boundaries of 'tubule'. SEM. x 4000. Fig. 27. Junction surface of enamel in area with no scallops. Note depression in end of prisms (arrow). SEM. x 3000.
scalloping when hard tissue formation commences. It seems likely that dimensional distortion occurring during maturation of the dentine and enamel (Marsland,1952) is the cause of scallop formation. Such a mechanism would explain the apparent disagreement between the findings of Schour (1960), who described a waviness in the free border of the ameloblasts, Provenza (1964), who could find no evidence for
Structure of the enamel-dentine junction
scalloping in the early stage of tooth formation, and Nylen & Scott (1960), who believed scallops to be formed at the time of mantle dentine formation. The present studies suggest that scallops do not form until after the first enamel is secreted. The dimensions of the scallops have been variously described in the past. Masukawa (1959) and Nakamura (1959) believed that they ranged between 3 and 4 ,csm. Nylen & Scott (1958) however recorded 100-700 ,um in both amptitude and frequency. The studies on mature teeth have confirmed the variable pattern of the junction not only in different teeth but within the same tooth. The similarity between the human deciduous and permanent teeth is remarkable in view of the environmental differences during development, but the shallower scallops in the former group may represent differences in maturation of the deciduous enamel and dentine resulting in less distortion. Little is known about the detailed differences, if they exist, between deciduous and permanent teeth. The variation in the pattern of the junction in human teeth requires further study since it is not clear why approximal areas should have more highly developed scallops than buccal and lingual dentine surfaces. Since lateral spread of caries occurs at the EDJ (Eccles & Green, 1973) it seems important to understand these structural differences which are perhaps produced by differential dimensional changes during maturation. The raised dense edges to the dentinal scallops could be due to distortion and relaxation of stress during decalcification, but this seems unlikely in view of the similarity of the shape and dimensions of scallops studied in undecalcified material. It seems more probable that the mantle dentine is supported by collagen fibres, and that weaker areas between them result in the formation of scallops as maturation of the dentine and enamel occurs. It may be that von Korff fibres, if they exist (Ten Cate et al. 1960; Whittaker & Adams, 1972), play some role in this process. The three dimensional arrangement of these fibres does not appear to have been studied. In the material sectioned through the junction the appearance of the zone of altered enamel was similar to that described previously using other methods (Helmcke, 1967). The prisms in altered enamel contain fewer crystallites, and tend to be fused together, so that their boundaries are indistinct. This zone is not always present, and further work is required to explain its significance. The dentinal tubules were clearly recognizable in the region of the junction, but branching, although present in this region, did not appear to be more than that seen in other parts of the dentine. The technique is not capable of preserving soft tissues, so that no comment can be made as to the presence or absence of odontoblast processes (Holland, 1976). However, there was no evidence of calcification of the tubules in the normal teeth examined (Tsatas & Frank, 1972). The enamel tufts present in the sectioned material presented an interesting feature in that many extended into, or had affected, the underlying dentine. This suggests that a stress release phenomenon might produce cracking in the newly formed dentine, as well as separating multiple groups of ameloblasts, resulting in the leaf-like pattern described by Osborn (1969). The clean separation of enamel from dentine, either by dehydration or by chemical techniques, suggests that there is less interdigitation of crystallites than was previously thought (Helmcke, 1953; Heuser, 1961; Nalbandian & Frank, 1962; Takuma, 1967; Watson & Avery, 1954). The surface coating and smooth contour of the enamel junction may indicate that the lamina densa of the developing tooth takes part in the formation of the junction itself. If a membrane remains, the dentinal appatite crystals presumably could not act as 'seeds' for calcification of the enamel, and another mechanism must be sought. The only previously published electron
D. K. WHITTAKER
micrograph of the junctional surface of the enamel appears to be a replica by Shroff (1966), and this also demonstrates a clean separation between enamel and dentine. The perforations visible in the enamel junction surface were of the order of size of enamel spindles, and if they do in fact represent channels for the odontoblast processes they appear to do so in two distinct ways. One type of spindle was at the crest of a convexity produced by the end of an enamel prism, and the spindle therefore was enclosed within a single prism. The second type of spindle lay between a group of prisms, and the walls of the defect were contributed to by a number of prisms. If newly formed enamel matrix is deposited around extensions of the odontoblast processes, then it is not surprising that the prism arrangement should vary in relation to the odontoblast processes. SUMMARY
The enamel-dentine junction of human and Macaca irus teeth has been studied using light and scanning and transmission electron microscopy. There are differences between species and within the same tooth. Scalloping, when present, is more developed in the cuspal areas and approximally, and the scallops appear to be formed at the time of maturation of the hard tissues. Enamel tufts and spindles are briefly discussed. The depth of scallops was less in the human deciduous than in the permanent teeth, but no differences were noted between monkey deciduous and permanent dentitions. I should like to express my thanks % Mr R. Watkins for technical assistance, and to Professor B. E. D. Cooke, in whose department this work was carried out. REFERENCES BRADLEY, D. E. (1957). Some carbon replica techniques for the electron microscopy of small specimens and fibres. British Journal of Applied Physiology 8, 150-161. ECCLES, J. D. & GREEN, R. M. (1973). The Conservation of Teeth, p. 5. London: Blackwell. FALIN, L. 1. (1961). Histological and histochemical studies of human teeth of the bronze and stone ages. Archives of Oral Biology 5, 5-13. GUSTAFSON, A. G. (1961). The histology of fluorosed teeth. Archives of Oral Biology 4, 67-69. HELMCKE, J. G. (1953). Atlas des menschlichen Zahnes im electronen mikroskopischen Bild. Part 1: Histologie des normalen Zahnes. Berlin: Transmare Phot. HELMCKE, J. G. (1967). In Structural and Chemical Organisation of Teeth (ed. A. E. W. Miles), Vol. 11, p. 160. New York: Academic Press. HEUSER, H. (1961). Die Strucktur des menschlichen Zahnschmelzes im oberflachenhistologischen Bild. (Replica-technik). Archives of Oral Biology 4, 50-58. HOLLAND, G. R. (1976). The extent of the odontoblast process in the cat. Journal of Anatomy 121, 133149. MARSLAND, E. A. (1952). A histological investigation of amelogenesis in rats. I1. Maturation. British Dental Journal 92, 109-119. MASUKAWA, K. (1959). Electron microscopic study on the ultrastructure of predentine. Bulletin of Department of Anatomy, Osaka Dental College, No. 20, 1-7. MEYER, W. (1925). Strittigie Fragen in der Histologie des Zahnschmelzes. Vierteljahrsschrift fur Zahnheilkunde 3, 3-9. MORTIMER, K. V. & TRANTER, T. C. (1971). A scanning electron microscope study of carious enamel. Caries Research 5, 240-263. NAKAMURA, M. (1959). Electron microscope researches on the ultrastructure of tooth germs. Journal of Electron Microscopy 8, 136-140. NALBANDIAN, J. & FRANK, R. (1962). Microscopie electronique des gaines des structures prismatiques et inter prismatiques de l'email foetal humain. Bulletin du Groupement internationale pour la recherche scientifique en stomatologie 5, 523-547. NYLEN, M. V. & ScoTT, D. B. (1958). An electron microscopic study of the early stages of dentinogenesis Public Health Services Publication Washington 1, 613.
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NYLEN, M. V. & ScoTT, D. B. (1960). Electron microscopic studies of odontogenesis. Journal of Indiana Dental Association 39, 406-421. ORBAN, B. (1929). Dental Histology and Embryology, 2nd ed., p. 45-48. Philadelphia: Blackiston's Son & Co. OSBORN, J. W. (1969). The three dimensional morphology of tufts in human enamel. Acta anatomica 73, 481-495. PROVENZA, D. V. (1964). Oral Histology, Inheritance and Development. p. 127, London: Pitman. RErrH, E. J. & BUTCHER, E. 0. (1967). In Structural and Chemical Organisation of Teeth (ed. A. E. W. Miles), vol. 1, p. 372. New York: Academic Press. RYWKIND, A. W. (1931). So-called scalloped appearance at the dentino-enamel junction. Journal of the American Dental Association 18, 1103-1110. SCHOUR, I. (1960). In Noyes Oral Histology and Embryology, 8th ed., p. 111. London: Kimpton. SCOTr, J. H. & SYMONS, N. B. B. (1974). Introduction to Dental Anatomy, 7th ed., p. 202. Edinburgh: Churchill Livingstone. SHROFF, F. R. (1966). Basic Dental Anatomy and Histology, p. 72. London: Kimpton. TAKUMA, S. (1967). In Structural and Chemical Organisation of Teeth (ed. A. E. W. Miles). vol. 1, p. 342. New York: Academic Press. TEN CATE, A. R., MELCHER, A. H., PuDy, G. & WAGNER, D. (1970). The non-fibrous nature of the von Korff fibres in developing dentine. A light and electron microscopic study. Anatomical Record 168, 491-524. TOMES, C. S. (1898). A Manual of Dental Anatomy. 5th ed., p. 34. London: Churchill. TSATAS, B. G. & FRANK, R. M. (1972). Ultrastructure of the dentinal tubule substar.ces near the dentinoenamel junction. Calcified Tissue Research 9, 238-242. TYLMAN, S. D. (1928). The dento-enamel junction. Journal of Dental Research 8, 615-622. WALKHOFF, 0. (1924). Normale Histologie menschlicher Zahne. Leipzig: Max Hesses. WATSON, M. L. & AVERY, J. K. (1954). The development of the hamster lower incisor as observed by electron microscopy. American Journal of Anatomy 95, 109-162. WHITTAKER, D. K. & ADAMS, D. (1972). Electron microscopic studies on Von Korff fibres in the human developing tooth. Anatomical Record 174, 175-190.