Transmission Electron Microscopy of Spheroids

Spheroid samples were harvested on day 5 time point and suspended in buffered glutaraldehyde solution as outlined in the previously published paper by Rhode et al. [37 (link)]. The spheroids were further fixed in 1% osmium tetroxide solution. Following this step, the spheroids were dehydrated using ethanol and propylene series and treated with Spurr’s resin and the blocks were polymerized. Then, 70 nm sample sections were generated and were imaged using FEI Tecnai 12 Bio-twin transmission electron microscope.

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Mukundan S., Singh P., Shah A., Kumar R., O’Neill K.C., Carter C.L., Russell D.G., Subbian S, & Parekkadan B. (2021). In Vitro Miniaturized Tuberculosis Spheroid Model. Biomedicines, 9(9), 1209.

Publication 2021

Tecnai 12 Bio-twin transmission electron microscope

Ethanol Glutaraldehyde Osmium tetroxide Propylene Spurr resin Transmission electron microscope Twin

Corresponding Organization : Rutgers, The State University of New Jersey

Other organizations : Center for Discovery, Hackensack Meridian Health, Cornell University

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Variable analysis

independent variables

Harvesting time point (day 5)

dependent variables

Ultrastructural characteristics of the spheroids

control variables

Buffered glutaraldehyde solution for initial fixation
1% osmium tetroxide solution for additional fixation
Ethanol and propylene series for dehydration
Spurr's resin for embedding and polymerization
Thickness of sample sections (70 nm)
Imaging method (FEI Tecnai 12 Bio-twin transmission electron microscope)

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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