„Supplement“:

only for those who are interested

Analytical electron microscopy

Element analysis / Element mapping

Element specific contrast,

Electron microscopic spectroscopy

The interaction of beam electrons with sample electrons can result in

― element specific energy loss of beam electrons and
― the emission of element specific X-rays

EFTEM: Energy-Filtered Transmission Electron Microscopy (images)

ESI: Electron spectroscopic Imaging
EELS/PEELS: Electron Energy Loss Spectroscopy (P= parallel detection)

EDX/EDS/EDXS: Energy Dispersive X-ray Spectroscopy

Energy Dispersive X-ray Microanalysis
……………..

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Energy dispersive X-ray spectroscopy (EDXS, EDX,
EDS, XEDS)
analysis (EDXA)
microanalysis (EDXMA)

Used mainly in SEM, STEM

Energy dispersive X-ray spectroscopy

EDX(S)

Electron beam

https://en.wikipedia.org/wiki/Energy-dispersive_X-ray_spectroscopy#/media/File:EDX-scheme.svg

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 EDX spectrum of Fe2O3

Iron oxide Fe2O3

- An e- of the inner shell of an atom is ejected into the vacuum.

- Vacancy is filled by an e- from a higher state.
- Excess energy is emitted as X-ray quantum.

https://de.wikipedia.org/wiki/Energiedispersive_R%C3%B6ntgenspektroskopie#/media/Datei:EDX-Spektrum.svg

EDX in biology

Plastic section of embedded rat pancreas

Nicole M. Pirozzi, Jacob P. Hoogenboom, Ben N. G. Giepmans (2018) Histochem Cell Biol 150; 509–520

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 - Energy-filtered TEM (EFTEM)
- Electron energy loss spectroscopy (EELS)
- Electron spectroscopic imaging (ESI)

Element specific energy loss

(ESI, EELS)
https://en.wikipedia.org/wiki/Energy-dispersive_X-ray_

Inelastically
spectroscopy#/media/File:EDX-scheme.svg

scattered

E0 - ΔE
Electron beam

Promoted to an
empty state

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 EFTEM (ESI/EELS): 2 techniques
Post-column filter In-column filter

https://eels.info/products/overview
Magnetic prisms separate elastically scattered e- from inelastically scattered e-

Electron spectroscopic imaging ESI (Omega filter)

Omega filter
(4 magnetic prisms)
separates inelastically
scattered e- from
elastically scattered e-
Slit Slit

Elastic imaging Inelastic imaging (ESI)

Energy-filtered image: Only inelastically scattered e- were used for imaging:
Slit removes inelastically scattered e- Slit removes elastically scattered e-
(improved image quality) = Elemental distribution map
Thin sections necessary (multiple scattering!)
Version 12-09-Z-EFTEMB c by Carl Zeiss SMT, Oberkochen
https://www.yumpu.com/en/document/read/3583777/in-column-eftem-technology-with-corrected-omega-filter-optronik

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 Electron spectroscopic imaging (ESI)

Ferritin ( Ø: 11 nm)
(Negative staining)
Fe

https://eels.info/about/techniques/eels-0
Apoferritin
Iron
core
Protein
©Y.-D. Stierhof

E = 0 eV E = 80 eV
above FeM2,3-edge

EELS (Post-column filter)

Spectrum
(of an image point)

Entrance aperture

Slit

Prism
Slit
Image

https://eels.info/about/techniques/eftem Gatan

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 EDX and EELS in biology
Plastic section of rat pancreas

SEM TEM

Nicole M. Pirozzi, Jacob P. Hoogenboom, Ben N. G. Giepmans (2018) Histochem Cell Biol 150; 509–520

EELS spectrum
Core level ionizations/Ionization edge:
Low loss excitations: Interaction with tightly
e- interact with weakly bound core electrons of the atoms
bound e- in the sample (= core-loss region),
differs for each element and shell

Plasmon peak:
Plasma resonance of
the valence electrons

Zero loss peak ZLP:

Without energy loss

https://eels.info/about/overview Gatan

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TEM combined with NanoSIMS (Nanoscale Ssecondary
Ion Mass Spectrometry)

First step:
- Imaging in a TEM
(e.g., a 100 nm section)

Second step:
- Ion beam erodes sample surface.
- Atomic collisions result in release of
secondary ions.
- A mass spectrometer measures
the mass and identifies it.
- The sample is scanned by the ion beam
→ map of elemental and isotope
distribution

SIMS is part of another talk

Nicole M. Pirozzi, Jacob P. Hoogenboom, Ben N. G. Giepmans (2018) Histochem Cell Biol 150; 509–520