This paper presents a detailed analysis to directly compare the morphology and chemistry of human tooth layers using advanced scanning electron microscopy (SEM) techniques together with supporting data from energy dispersive spectroscopy (EDS) measurements. The aim of this study was to visualise and evaluate the structural and microanalytical differences of the mineralised hard tissues of human teeth. The extracted sound teeth without any pathologies were divided into the following groups: incisors, canines, premolars, and molars. Tooth samples were broken vertically to preserve the primary structures and to visualise individual tooth tissues. Specimens were also used to find variations in the elemental composition of tissues for different tooth groups. The average thickness of the enamel in the tooth groups studied was 1.1 mm and the average width of the enamel prisms was 4.2 µm, with the highest values observed for molars. The analysis of the chemical composition of the enamel showed that Ca and P were among the predominant elements. The average dentine thickness was 1.87 mm, with the highest values determined for molars, and the lowest for canines. The width of the dentinal tubules was less than 2 µm, for molars being significantly smaller. The analysis of the chemical composition of the dentine showed the highest O content of the all tooth tissues analyzed, while a lower P and Ca content was observed compared to the enamel. The cementum thickness averaged 0.14 mm, with the highest values observed for molars and the lowest for incisors. The analysis of the chemical composition of the cementum showed the lowest average O and P content, and the highest average C and N content, compared to the enamel and the dentine. Increasingly accurate imaging and analysis of dental hard tissue structures provides the opportunity for multifactorial evaluation in terms of their clinical application.