Oct 11, 2022

Public workspaceSample preconditioning before scanning electron microscopy (SEM)

DOI
https://dx.doi.org/10.17504/protocols.io.36wgqj465vk5/v1
  • An.Huang 1
  • 1XJTLU
An.Huang
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DOI: https://dx.doi.org/10.17504/protocols.io.36wgqj465vk5/v1
Protocol Citation: An.Huang 2022. Sample preconditioning before scanning electron microscopy (SEM). protocols.io https://dx.doi.org/10.17504/protocols.io.36wgqj465vk5/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: October 10, 2022
Last Modified: October 11, 2022
Protocol Integer ID: 71066
Keywords: preconditioning method, scanning electron microscopy, bacterial sample, scanning electron microscope, electron microscopy, methods of bacteria, electron microscope, microscopy, bacteria, sem
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Abstract
Bacterial samples need to be preconditioned before visualized by scanning electron microscope (SEM). This protocol describes preconditioning methods of bacteria before SEM.
Troubleshooting
Bacterial samples preconditioning
10m
Obtain a small quantity of bacteria to be tested from a culture.
Centrifuge the samples atCentrifigation5000 rpm, 4°C, 00:10:00 . Resuspend the pellet using 1X phosphate buffed saline (PBS). Repeat this step for three times. Keep the supernatant after the third centrifuge and preserve the samples at Temperature4 °C .

10m
SEM sample preconditioning
2h
Wash silica wafers with ddH2O and acetone for three times.
Add samples onto the silica wafer using a pipette tip.
Incubate the silica wafers in fixation solution (2.5% glutaraldehyde in 1X PBS) atTemperature4 °C overnight.

Incubate the samples in ethanol solution with an increasing concentration (40%, 70%, 96%, 100%) for Duration01:00:00 each concentration.

1h
Dry the samples at room temperature for Duration01:00:00 .

1h
Use a precision etching coating system, sputtering the samples with Au-Pd alloy.

Note
If samples contain gold, sputter the samples with chromium.

The samples are ready to be visualized using SEM