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. 2019 Nov 26;9:17625. doi: 10.1038/s41598-019-53875-y

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© The Author(s) 2019

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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Comparison between measured and modelled PSFs of a light sheet microscope. (a1) 200 nm sized fluorescent beads were recorded using a 10x objective with NA = 0.3 (UPLFLN 10x, Olympus, Germany). The figure shows a maximum intensity (MIP) projection (xz-direction) obtained from 402 slices. (a2) PSF extracted by registration and averaging of 10 manually selected beads from (a1) (left), compared to a calculated PSF according to Eq. (15) (Model parameters: NA = 0.3, λ_ex = 488 nm, λ_em = 520 nm, n = 1.561, f = 80 mm, d = 8 mm, no damping). (a3) comparison of lateral (x, y = 0, z = 0) and axial (x = 0, y = 0, z) intensity profiles of the measured and the modelled PSFs depicted in (a2). (b1) 200 nm fluorescent beads were recorded using a 20x objective with NA = 0.45 (LUCPLFLN, Olympus, Germany). The panel shows a maximum intensity projection (MIP, xz-direction) of the 3D reconstructed fluorescent beads obtained from 500 slices. (b2) PSF obtained after registration and averaging of 10 manually selected beads from b1 (left) versus a calculated PSF according to Eq. (15). (Model parameters: NA = 0.6, λ_ex = 488 nm, λ_em = 520 nm, n = 1.561, f = 80 mm, d = 8 mm, no damping). (b3) comparison of lateral (x, y = 0, z = 0) and axial (x = 0, y = 0, z) intensity profiles of the measured and of the modelled PSFs depicted in (b2). As for the 10x objective visual comparison confirms a good agreement between measurements and theory.