The compound eyes of trilobites, which appeared in the Early Cambrian, represent one of the first preserved visual systems. Application of state-of-the-art microscopy techniques in the present study has revealed fine details of the microstructure and chemistry of these unusual calcite eyes that, until now, have been inaccessible and this has facilitated new insights into their growth and function. Six species from three families of trilobite with holochroal eyes, ranging from Early Ordovician to Middle Carboniferous, and 21 species from three families of trilobite with schizochroal eyes, ranging from Early Ordovician to Middle Devonian, were investigated. High-resolution microscopy techniques, including Electron Backscatter Diffraction, Transmission Electron Microscopy and Electron Probe Micro-analysis have made it possible to ‘see’ through the diagenesis of trilobite lenses to reveal the likely original lens microstructure and chemistry. Computer-based optical modelling has further shown how original lens microstructure and chemistry enhanced lens function. The discovery of sub-micron sized crystals that display a gradual and precise change in orientation shows that in many lenses much of the original microstructure has remained intact despite exposure to pore fluids and elevated temperatures and pressures during diagenesis. Although microstructure varies slightly with lens shape, there is often exceptionally precise crystal orientation. In holochroal eyes there is a direct relationship between lens shape and microstructure; where lens surfaces are planar, crystals are of uniform orientation, but where lens surfaces are convex, c axis orientation fans out, away from the lens axis. Several microstructural patterns have been identified in schizochroal lenses. However, a single original microstructural pattern, in which c axis orientation fans out at lens surfaces but remains parallel to the lens axis in the centre of the lens, may be applicable to all schizochroal lenses. The original chemical composition of the lenses, in particular those of schizochroal type, is less commonly preserved. However, the unravelling of a diagenetic pathway of change, through understanding the intricacies of relationships between different minerals in the lenses, has made possible a better understanding of how these lenses were altered during diagenesis. Lenses in holochroal eyes are invariably low-magnesium calcite, like the rest of the exoskeleton, as has been established for some time. The present study clarifies the original structures of schizochroal lenses and in doing so, ends the controversy over lens function; lenses were originally constructed as doublets, as suggested by Clarkson and Levi-Setti, and were not gradient index lenses, as was suggested by Campbell and Bruton and Haas. Lenses in schizochroal eyes, were constructed of high-magnesium calcite, with highest concentrations of magnesium in the lower ‘intralensar bowl’ and central ‘core’ regions of the lens. The degree of partitioning of high levels of magnesium, of up to 8 mole % MgCO3, in the schizochroal eyes is remarkable given the magnesium-poor ‘calcite seas’ in which they were formed. This is perhaps the first example of element partitioning within biominerals for a specific function. Based on the growth sequence of modern arthropod exoskeletons lenses in trilobite eyes are likely to have grown from the outer surface in, one lamella at a time, with microstructure and chemical composition controlled by an organic matrix. Assessment of the trilobite optical structures, using Code V optical modelling software, leads to the conclusions that the trilobite eyes functioned in a similar manner to the apposition eyes of modern animals. Code V modelling of holochroal and schizochroal eyes, and the subsequent determination of their resolution and sensitivity, shows that both eye types probably had a single optically isolated photoreceptor beneath each lens. Using a combination of specific lens size, shape, spacing, microstructure and composition, the schizochroal eye was adapted to low light intensities, similar to the eye of the modern isopod Cirolana. These adaptations would have provided the trilobite with light and dark detection of a resolution sufficient to identify movement, allowing it to detect prey and defend itself against predators. The birefringent properties of the calcite from which these lenses were made could be a hindrance, resulting in double refraction of light rays and the formation of ‘ghost’ images. Fascinatingly however this property provides the lenses with the refractive power required to make full use of the light available to them, vital for an organism with a crystalline lens with a fixed focal length. Study of the calcified lenses of ostracods and brittlestars and comparison to lenses in schizochroal trilobite eyes confirms that these modern organisms do not provide accurate analogues for trilobite eyes. No other organism that shares all characteristics of schizochroal trilobite eyes has yet been found; the eyes of the phacopine trilobites remain unique in the natural world.