Data underlying the publication: Depth-dependent scaling of axial distances in light microscopy

IntroductionData underlying the publication: Depth-dependent scaling of axial distances in light microscopyPublication DOI: https://doi.org/10.1364/OPTICA.520595Repository created by Daan Boltje<br>AbstractIn volume fluorescence microscopy, refractive index matching is essential to minimize aberrations. There are however, common imaging scenarios, where a refractive index mismatch (RIM) between immersion and sample medium cannot be avoided. This RIM leads to an axial deformation in the acquired image data. Over the years, different axial scaling factors have been proposed to correct for this deformation. While some reports have suggested a depth-dependent axial deformation, so far none of the scaling theories has accounted for a depth-dependent, non-linear scaling. Here, we derive an analytical theory based on determining the leading constructive interference band in the objective lens pupil under RIM. We then use this to calculate a depth-dependent re-scaling factor as a function of the numerical aperture (NA), the refractive indices 𝑛₁ and 𝑛₂, and the wavelength 𝜆. We compare our theoretical results with wave-optics calculations and experimental results obtained using a novel measurement scheme for different values of NA and RIM. As a benchmark, we recorded multiple datasets in different RIM conditions, and corrected these using our depth-dependent axial scaling theory. Finally, we present an online web applet that visualizes the depth-dependent axial re-scaling for specific optical setups. In addition, we provide software which will help microscopists to correctly re-scale the axial dimension in their imaging data when working under RIM.<br>General descriptionAll data underlying the manuscript figures are stored in this repository. Experiments have been carried out at the Department of Imaging Physics at Delft University of Technology (Delft, NL) between December 2022 and February 2023.<br>The repository contains images generated by wave-optics calculations, images acquired on a confocal microscope imaging under RIM and image z-stacks of fluorecent beads residing on the coverslip, and sapphire ball. Such measurement is done for different numerical aperture objectives and refractive indices. Refer to the README for more information.<br>

## 论文关联数据
本数据集支撑的发表论文：光学显微镜中轴向距离的深度依赖缩放关系
论文DOI：https://doi.org/10.1364/OPTICA.520595
本数据集仓库由Daan Boltje创建

## 摘要
在体积荧光显微镜成像中，折射率匹配是抑制像差的必要手段。但在多数常见成像场景中，浸液介质与样品介质间的折射率失配（Refractive Index Mismatch, RIM）难以避免。该折射率失配会导致采集的图像数据产生轴向形变。多年来，学界已提出多种轴向缩放因子以校正此类形变。尽管已有研究指出轴向形变存在深度依赖性，但迄今为止所有缩放理论均未考虑深度依赖的非线性缩放问题。本文基于折射率失配条件下物镜光瞳内的主导相长干涉带推导得到解析理论，并据此计算得到以数值孔径（Numerical Aperture, NA）、折射率n₁与n₂以及波长λ为变量的深度依赖重缩放因子。我们将理论计算结果与波动光学计算结果，以及采用新型测量方案获得的不同数值孔径与折射率失配条件下的实验结果进行对比。作为验证基准，我们在不同折射率失配条件下采集了多组数据集，并利用本文提出的深度依赖轴向缩放理论对其进行校正。最后，我们开发了一款在线网页小程序，可针对特定光学系统可视化深度依赖的轴向重缩放效果。此外，我们还提供了配套软件，可帮助显微镜工作者在存在折射率失配的成像场景下，对采集数据的轴向维度进行准确的重缩放处理。

## 通用说明
本仓库存储了支撑论文所有配图的全部实验数据。实验于2022年12月至2023年2月间在荷兰代尔夫特理工大学成像物理系完成。
仓库包含波动光学计算生成的图像、存在折射率失配条件下共聚焦显微镜采集的图像，以及位于盖玻片与蓝宝石球上的荧光微球的图像Z堆叠数据。上述测量针对不同数值孔径的物镜与不同折射率条件完成。更多细节请参阅README文件。