The ability of cornea to transmit light whilst being mechanically resilient is directly attributable to the formation of an extracellular matrix containing orthogonally arranged sheets of densely packed uniformly-narrow collagen fibrils, in which each sheet contains parallel arrays of fibrils. An understanding of the molecular basis of tissue organisation requires knowledge of the detailed 3-dimensional structure of individual collagen fibrils and how they interact to generate higher order structures. in this study adult bovine corneal collagen fibrils were examined using transmission electron microscopy and automated electron tomography to generate three-dimensional volumes of individual fibrils which had been negatively stained. Tilt series images were collected with an angular increment of 2° over a range of ± 70° using a Philips CM200FEG coupled to a TVIPS automated electron tomography system. A microfibrillar substructure was observed throughout the thickness of the fibril (Figure 1). The microfibrils were ∼4 nm in diameter and were oriented in a right-handed helical twist along the length of fibrils. The microfibrils were observed at an angle of 15° to the long axis of the fibril. As the technique of electron tomography generates a threedimensional volume from the data it was possible, for the first time, to visualise the lateral arrangement of molecules within individual fibrils. There were regions where there was a distinct lateral order and other regions which showed a more fluid-like arrangement. The axial positions of most crystallinity corresponded with the location of the C-telopeptides and N-telopeptides as well as a region close to the d-band in the gap structure of the fibrils. The study also demonstrated that the microfibrils are organised in a quasi-hexagonal packing arrangement