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High-Resolution Imaging of Tumor Spheroids and Organoids Enabled by Expansion Microscopy.

Frontiers in molecular biosciences (2020-11-17)
Steven J Edwards, Valentina Carannante, Kyra Kuhnigk, Henrik Ring, Tatsiana Tararuk, Finn Hallböök, Hans Blom, Björn Önfelt, Hjalmar Brismar

Three-dimensional cell cultures are able to better mimic the physiology and cellular environments found in tissues in vivo compared to cells grown in two dimensions. In order to study the structure and function of cells in 3-D cultures, light microscopy is frequently used. The preparation of 3-D cell cultures for light microscopy is often destructive, including physical sectioning of the samples, which can result in the loss of 3-D information. In order to probe the structure of 3-D cell cultures at high resolution, we have explored the use of expansion microscopy and compared it to a simple immersion clearing protocol. We provide a practical method for the study of spheroids, organoids and tumor-infiltrating immune cells at high resolution without the loss of spatial organization. Expanded samples are highly transparent, enabling high-resolution imaging over extended volumes by significantly reducing light scatter and absorption. In addition, the hydrogel-like nature of expanded samples enables homogenous antibody labeling of dense epitopes throughout the sample volume. The improved labeling and image quality achieved in expanded samples revealed details in the center of the organoid which were previously only observable following serial sectioning. In comparison to chemically cleared spheroids, the improved signal-to-background ratio of expanded samples greatly improved subsequent methods for image segmentation and analysis.

Product Number
Product Description

Minimum Essential Medium Eagle, With Earle′s salts and sodium bicarbonate, without L-glutamine, liquid, sterile-filtered, suitable for cell culture
Heparin sodium salt from porcine intestinal mucosa, Grade I-A, ≥180 USP units/mg
Taurine, ≥99%
(−)-Blebbistatin, solid, synthetic
Collagenase from Clostridium histolyticum, Sigma Blend Type F, ≥2.0 FALGPA units/mg solid