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Reference isotropic 3D electron microscopy data of SARS-COV-2-infected grivet kidney epithelial (Vero) cells at 24 h after exposure

posted on 2021-10-04, 21:42 authored by Song Pang, John Bernbaum, Jens H Kuhn, Harald HessHarald Hess, C. Shan XuC. Shan Xu

Understanding cellular architecture is essential for understanding biology. Electron microscopy (EM) uniquely visualizes cellular structure with nanometer resolution. However, traditional methods, such as thin-section EM or EM tomography, have limitations because they only visualize a single slice or a relatively small volume of the cell, respectively. We overcame these limitations by imaging whole cells and tissues via the enhanced focused ion beam scanning EM (FIB-SEM) platform at 4-nm voxels with month-long acquisition. We used this approach to generate diverse reference 3D image datasets and hope to create a reference library to explore comprehensive quantification of whole cells and all their constituents. These open-access data offer a foundation upon which to nucleate a new volume of high-resolution whole-cell EM images and subsequent analyses, a new paradigm that invites biologists to explore and pose questions with a fresh perspective.

SARS-CoV-2-infected grivet kidney epithelial (Vero) cells at the 24-h time point show a high rate of infection with visible convoluted membranes, double-membrane vesicles, vesicle packets, and virus particles.

Sample: SARS-COV-2-infected grivet kidney epithelial (Vero) cells at the 24-h time point

Protocol: CCL-81 cells were seeded in a T75 flask at 80% confluency and infected with SARS-CoV-2, at a MOI of 0.01. After 24 hours, the cells were collected from flasks and preserved in 2.5% Glutaraldehyde (E.M. Sciences, Warrington, PA), in Millonig’s Sodium Phosphate Buffer (Tousimis Research, Rockville, MD). After fixation was complete, the cells were washed repeatedly in Millonig’s Buffer, and incubated for two hours in 1.0% Osmium Tetroxide, in the same buffer. Following rinsing steps in Ultrapure Water and en bloc staining with 2.0% Uranyl Acetate, the samples were dehydrated in a series of graded ethanols, and infiltrated and embedded in Spurr’s plastic resin (E.M. Sciences).

Contributions: Sample provided by John Bernbaum and Jens H. Kuhn ( NIH/NIAID/DCR/Integrated Research Facility at Fort Detrick), prepared for imaging by Song Pang (HHMI/Janelia), imaging by Song Pang (HHMI/Janelia), and post-processing by C. Shan Xu (HHMI/Janelia)

Dataset ID: jrc_ccl81-covid-1

Final voxel size (nm): 4.00 x 4.00 x 4.00 (X, Y, Z)

Dimensions (µm): 15 x 20 x 25.8 (X, Y, Z)

Acquisition date: 2020-09-10

Dataset URL: https://data.janelia.org/FBPYq

Visualization Website: https://openorganelle.janelia.org/datasets/jrc_ccl81-covid-1