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Electron tomographic reconstruction of plastic-embedded organelles involved in the chitin secretion process

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Shillito, B., Koster, A. J., Walz, J., & Baumeister, W. (1996). Electron tomographic reconstruction of plastic-embedded organelles involved in the chitin secretion process. Biology of the Cell, 88(1-2), 5-13.

Structural investigations on the chitin secretion system of a deep-sea hydrothermal vent tubeworm, Riftia pachyptila, are presented. In this organism, 300-nm cup-shaped organelles are responsible for the formation of large (50 nm) crystalline chitin microfibrils of the rare beta form. Electron tomographic reconstructions of plastic-embedded cup-shaped organelles were carried out, using 93 projections distributed over a +/- 70 degrees angular range with a 0.63-nm pixel size. In addition, the shrinkage profile of the plastic sections was measured to determine the required imaging conditions. The entire data collection was done automatically and carried out under cryo-conditions. Automation resulted in an efficient use of irradiation dose; 96% of the total dose is required for the data itself, only 4% is used for the required compensation of focus change and image shift. The results are presented as 3-nm thick slices through the 3-D reconstruction, (parallel to the original thin section, ie the x-y plane). The 3-nm slices allow to visualize separate structures that are overlapping in a conventional 2-D projection of the initial thin section. Also, these thin slices through the 3-D reconstruction show the fine contours of the nascent microfibril tip, which appears to be cone-shaped. [References: 33]