Louis-Marie Lebas (Villeurbanne / FR), Abid Zulfiqar (Saarbrücken / DE), Akkiz Bekel (Villeurbanne / FR), Lucian Roiban (Villeurbanne / FR), Karine Masenelli-Varlot (Villeurbanne / FR)
Abstract text (incl. figure legends and references)
Environmental electron microscopy has been developed to study specimens under water or gas partial pressure [1]. It allows the preservation of hydrated samples with minimum preparation. In ESEM, the spatial resolution can reach the nanometer, and in ETEM the Ångström. Using a specific tomographic sample holder, tilt series can be recorded and reconstructed in a 3D volume [2, 3].
This work aims to establish liquid-phase and three-dimensional scanning electron microscopy (LPSTEM) as a new technique for characterizing biological or materials science samples at the nanoscale, by addressing two of the remaining limitations. The electron dose should be reduced and quantified, especially for sensitive materials. The acquisition time should be commensurate with the events to be studied, i.e. no more than few minutes in most of operando experiments.
We show here automated acquisitions of ± 70° tilt series in LPSTEM in less than 5 minutes in ESEM and ETEM. SE, BF and HAADF images are simultaneously recorded. Moreover, the electron dose can be precisely controlled. We use a ThermoFisher (FEI) QuattroS ESEM operated at 30 kV and a ThermoFisher (FEI) Titan 80-300 ETEM at 300 kV.
Researches are conducted on SBA latex and on different types of gels: silica aerogels and different types of aluminium hydroxide wet gel suspensions. The specimen are deposited on a TEM grid coated with a holey carbon film, itself fixed on a home-made prototype of wet-STEM tomography sample holder (ESEM), or on a dedicated sample holder (ETEM) [4]. Images are recorded with our home-made software M-SIS based on Python, using Autoscript and Digital Micrograph libraries. It features automatic eucentric, drift correction and various helps for automatic acquisition. The tilt series are aligned with the TomoJ and data are segmented with 3D Slicer.
The 3D models exhibit a spatial resolution around 10 nm (ESEM) or around few nanometers (ETEM) whereas the electron dose can be finely controlled and can be kept below the viability threshold of a biological cell. Hence the morphology of the samples and their porosity are studied. The limits will also be exposed in term of sample thickness, liquid film thickness control, spatial resolution, electron dose and total imaging time.
These results open the way to perform fast multiscale studies in liquid-phase STEM tomography with easy sample preparation, both in ESEM and ETEM. This low-dose process could be applied for various types of hydrated or liquid samples and could be widely used in materials characterization.
[1] Schuh, T., de Jonge, N. (2014) "Liquid scanning transmission electron microscopy: Nanoscale imaging in micrometers-thick liquids" Comptes Rendus Physique, 15(2-3)
[2] Masenelli-Varlot, K. et al. (2014) "Wet-STEM Tomography: Principles, Potentialities and Limitations", Microscopy and Microanalysis, 20(2).
[3] de Jonge, N. (2018) "Theory of the spatial resolution of (scanning) transmission electron microscopy in liquid water or ice layers", Ultramicroscopy, 187.
[4] Jiao, X. et al. (2019) "Electron tomography on latex particles suspended in water using environmental scanning electron microscopy", Micron, 117.
Figure 1: From upper-left to bottom-right: YZ, XY and XZ ortho-slices of the reconstructed silica airgel volume from images
recorded at different tilt angles a) in BF mode, b) in HAADF mode. Scale bar is 500 nm.