TEM micrographs from human pancreatic islets derived from stem cells

TEM micrographs from human pancreatic islets derived from stem cells

Description

Samples from SC-islets and human islets were chemically fixed with 2.5% glutaraldehyde in 0.1 M sodium cacodylate buffer, pH 7.4, supplemented with 2 mM calcium chloride at RT, for 2 h. After washing, the specimens were osmicated in the same buffer with 1% nonreduced osmium tetroxide on ice, for 1 h. Specimens were then washed and dehydrated in increasing concentration of ethanol and acetone, before gradual embedding into Epon. After polymerization over 18 h at 60 °C, a pyramid was trimmed on the location of the embedded cells. Ultrathin, 60-nm sections were cut using an ultramicrotome, picked on Pioloform-coated single-slot grids and poststained with uranyl acetate and lead citrate. Micrographs were acquired with a Hitachi HT7800 microscope operated at 100 kV using a Rio9 CMOS-camera. One or two SC-islets were chosen randomly for examination and 11–12 SC-islet beta cells were selected for imaging on the basis of characteristic features of beta-like granules. Cells for analysis from two differentiation experiments were selected based on the characteristics features of mature granules and images for α-cells and β-cells were collected with random systematic sampling: 3-5 images per cell, 4 cells per aggregate and 5 aggregates per sample. ER morphology was evaluated from the micrographs acquired at nominal magnification of 5,000X.
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Year of publication

2022

Authors

Helena Vihinen Orcid -palvelun logo - Creator

Eija Jokitalo Orcid -palvelun logo - Curator

Other information

Fields of science

Biochemistry, cell and molecular biology

Language

English

Open access

Restricted access

License

Creative Commons Attribution 4.0 International (CC BY 4.0)

Keywords

Electron microscopy, transmission electron microscopy

Subject headings

elektronimikroskopia
TEM micrographs from human pancreatic islets derived from stem cells - Research.fi