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. 2021 Mar 30;12:1957. doi: 10.1038/s41467-021-22251-8

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — a, b Comparison of the target displacement during tilting assuming that the sample is flat compared to the center of rotation (a, green arc) or when considering an off-plane shift (b, red arc and red lines). Note that the trajectory of an off-plane target is different than for an eucentric target. c Measurement of the median displacement in the x-coordinate (blue) and y-coordinate (red) without correcting for off-plane shift (left panel) and with correction (right-panel) over multiple tilt-series (n=150). Error bars represent the inter quartile range (IQR). d Measurement of the median defocus variation from the first image without correcting for off-plane shift (left panel) or with correction (right-panel) over multiple tilt-series (n = 150). Error bars represent the IQR. In general, errors in defocus originate mainly from errors in targeting and are not systematic.