Beyond the naked

eye: Characterizing
nanomaterials
with precision
Today’s demand for faster
and smaller electronics is
ever growing. Discover how
to meet this demand using
the latest nanomaterial
characterization techniques

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Characterizing nanomaterials

Introduction
As the demand for faster, smaller, and more
cost-effective electronics remains high, so
does the need for researchers to innovate and
develop novel solutions that deliver superior
performance, speed, and reliability.
These innovations are driving the
development of new and exciting
technologies, and part of this change is
the ability for users to characterize
materials with more precision than ever
before. By characterizing the properties
of a material at the micro and nanoscale,
researchers gain valuable insights into the
fundamental behavior of a material, leading
to new discoveries and breakthroughs.
Nevertheless, the research can only be as
good as the microscope and instrument
used for the experiment.
It’s vital that scientists select the most
appropriate instrument for their needs - to
ensure they can precisely and cost-effectively
characterize samples. The instrument
selection process often involves choosing
an instrument that offers the sensitivity and
resolution they require for sample analysis.
Plus, the instrument must be easy-to-operate
and maintain, with minimal downtime. For
Contents
• Exploring the microscopic world with
3D X-ray imaging
• Observation of mechanical testing at
the nanoscale
• Unlocking the microscopic mysteries
of 3D printed powders
• ZEISS Crossbeam 550: The future of
high-throughput imaging
• The power of laser: Optimizing TEM
sample preparation
• Seeing beyond the surface: Exploring
crystallographic information with
diffraction contrast tomography
• Advancing nanotomography with
X-ray technology
• Next-generation electron optics:
Taking imaging to new heights
• Unlocking perovskite catalysts with
sweet spot imaging
• Featured products

2
Characterizing nanomaterials
instance, material scientists need to be able
to explore a material in as much detail as
possible, locate and investigate regions of
interest within a large sample, and effectively
characterize large three-dimensional volumes.
This application eBook presents a guide
based on some of the most advanced material
characterization technologies on the market.
This eBook will explain how to investigate the
microscopic world using 3D X-ray imaging,
explore how to fully characterize powders,
and highlight the future of high-throughput
imaging using focused ion beam-scanning
electron microscopy (FIB-SEM) systems.
Plus, we will show users how to optimize
sample preparation, explore crystallographic
information using diffraction contrast
tomography, and much more. This expert
guide will also introduce the latest
cutting-edge technologies from ZEISS – one
of the leading providers of microscopic and
characterization tools. These tools have
been designed to support both academic
researchers and industries in materials
science by delivering the most advanced
electron microscopy (SEM), FIB-SEM, and
X-ray microscopy (XRM) tools, among
many others.
Exploring the microscopic world with
3D X-ray imaging
Micro- and nano-computer tomography
(micro and nano-CT) are powerful and
non-destructive imaging techniques that use
X-rays to generate high-resolution 3D images.
There have been many significant advances in
both micro-CT and nano-CT technology, which
have resulted in improved resolution, faster
scan times, and greater sensitivity levels.
These technological advances have enabled
researchers to capture high-resolution images
like never before, with increased throughput
and efficiency. Another key advancement is 3D
X-ray microscopy (XRM), which delivers even
higher resolution levels. In this application
note, learn how to objectively evaluate the
resolution performance of XRM and other
CT instruments relevant to a users research
and production goals. Plus, learn how spatial
resolution delivers researchers with the
most meaningful method for evaluating an
instrument’s performance, and how XRM
instruments from ZEISS have proven to
deliver the highest spatial resolution across
the widest range of working distances.
For more information on XRM, take a look
at this resource - which explores mechanical
testing at the nanoscale. This application
note introduces in situ mechanical testing
integrated with nanoscale XRM (which holds
great promise for a range of applications, from
metals and biomaterials to thin coatings).
Plus, be introduced to the Xradia Ultra Load
Stage, a new nanomechanical test stage that
is available for integration into the ZEISS
Xradia Ultra platform, the only nanoscale
XRM available for the laboratory.
Unlocking the microscopic mysteries of
3D printed powders
The bulk density and flow behavior are
important parameters for characterizing
powders in powder technology - since they
can directly impact the efficiency and quality
of the final powered product.
Bulk density refers to the mass of a powder
per unit volume, which can affect the packing
and flow behavior of a powder. In contrast,
flow behavior refers to the way a powder
moves and behaves when subjected to
external forces, such as mechanical pressure
- which can affect their processing and
handling. To explore this further, look at this
application note to find out more about these
vital parameters in detail. This resource will
also explain how light and scanning electron
microscopy combined with an image analysis
tool makes it possible to fully characterize
powders. You’ll find out how this powerful
tool allows for automatic segmentation
and analysis of particles, pores, and phases
through gray level thresholding to allow
users to obtain quick and reliable statistical
information about the quality of a powder.
ZEISS Crossbeam 550: The future of
high-throughput imaging
A FIB-SEM is an advanced microscopy
instrument that combines two powerful
techniques together and delivers
high-resolution imaging, along with precise
material analysis. The FIB-SEM system
comprises of two key components: the
focused ion beam (FIB) and the scanning
3
Characterizing nanomaterials
electron microscope (SEM). The FIB
uses a beam of charged ions to etch a
sample’s surface, while the SEM uses a
beam of electrons to scan the sample’s
surface to create high-resolution images
of its topography.
ZEISS is a leading manufacturer of
cutting-edge microscopy solutions,
including the latest FIB-SEM systems.
ZEISS and its team of experts have made
significant advancements to its product
portfolio by improving image resolution,
contrast, ease-of-use, and throughput
offered by its instruments. As a result,
the new ZEISS Crossbeam 550 has
become the go to instrument of choice
for high-throughput FIB-SEM work. If
you’re interested in learning more about
the capabilities of this instrument, take
a look at this application note. This
resource provides valuable insights into
the performance of the Crossbeam 550
in the lab and how it can deliver a high
imaging resolution for SEM beam currents
up to several nA.
The power of laser: Optimizing TEM
sample preparation
LaserFIB instruments have gained huge
popularity over recent years, due to their
powerful capabilities in the field of materials
science. Their growing popularity is mostly
because of their high precision, fast speed,
high versatility, and non-destructive nature.
This technique uses a combination of laser
and ion beam technology to either alter
or analyze a given material at the micro
or nanoscale. LaserFIB achieves this by
combining a pulsed laser with a focused ion
beam, such as gallium ions, to deliver precise
material removal or modification.
In this resource, find out more about
LaserFIB and the value of femtosecond
(fs) laser ablation for site-specific
transmission electron microscopy (TEM)
sample preparation. This resource will also
introduce the ‘Cut-to-ROI’ workflow. This is
a specialized method developed to enable
users to explore a region of interest that
is located deep within a given sample and
unable to be assessed using direct FIB
instruments. Plus, this resource will introduce
the ZEISS Crossbeam laser system and how
the instrument allows for 3D-site-specific
preparation of deeply buried regions of
interest, and the preparation of multiple
TEM-suitable windows spread over a large
sample area.
Seeing beyond the surface: Exploring
crystallographic information with diffraction
contrast tomography
X-ray tomography is a powerful technique
that uses X-rays to generate detailed, 3D
images of a sample. The X-ray source emits
a strong beam of X-rays that are either
attenuated or absorbed to different degrees,
depending on the composition of a material.
The detector records the intensity of the
X-rays, which is used to non-destructively
generate a detailed 3D image of a sample.
In this resource, find out more about this
technique and the two popular contrast
mechanisms used with X-ray tomography,
termed X-ray absorption and phase contrast.
This application note will also introduce the
Synchrotron-based XRM instrument, which
has been designed to deliver crystallographic/
diffraction information from poly-crystalline
samples and demonstrate how the Versa
series of X-ray microscopes can help support
such imaging goals.
Plus, click here to find out more about
X-ray microscopy and how this technique
can enable users to non-destructively
investigate a range of samples, and how
the Xradia Ultra (the only available
laboratory-based nanoscale X-ray
microscope), achieved resolution down
to 50 nm using X-ray focusing.
Next-generation electron optics: Taking
imaging to new heights
SEM has advanced significantly over recent
years, with improvements in imaging
resolution, speed, and sample preparation
techniques. Technological advances have
enabled us to explore materials like never
before. Many new and advanced instruments
now come equipped with additional detectors
and automated imaging capabilities, which
enable users to rapidly image samples with
high accuracy and reproducibility. Key to the
performance of any SEM, conventional or
4
Characterizing nanomaterials
field emission SEM (FE-SEM), is its electron
optical column. In this resource, find out
how ZEISS contributed directly to the field
by developing the Gemini electron optics,
the Nano-twin lens, and their latest
innovation - the EO optical engine. Find
out how the EO engine delivers increased
resolution at low kV by optimizing the
convergence angle of electrons travelling
through a column, and also increases
the available field of view using the
high-performance Nano-twin lens.
Unlocking perovskite catalysts with sweet
spot imaging
Platinum (Pt) is one of the rarest, yet most
versatile metals to exist. Pt is used within
a whole range of industrial and commercial
applications, from jewellery and lab equipment
to electronics and engineering. Pt is now
widely used for catalytic chemical reactions
in automobile, chemical refining, and energy
industries. To determine the catalytic ability of
platinum nanoparticles, it’s essential that we
understand their structure. Undesirably, when
using conventional SEM methods they do not
reveal enough comprehensive morphological
information about these particles due to the
lack of electron beam stability. Fortunately,
there are techniques available to accurately
determine particle size and morphology.
In this application note, find out more about
sweet spot imaging conditions and how they
can be determined using the advantages
of the electron optics of the ZEISS FE-SEM
range. Plus, learn how to find the optimal
conditions for imaging a range of samples
and how the stability and stage of the
ZEISS GeminiSEM family were found to
increase image stabilization and brilliance,
resulting in high resolution imaging at low
kilo-electron-volt (keV).
5
Characterizing nanomaterials

Technical Note

Resolution of a 3D X-ray Microscope

Defining Meaningful Resolution Parameters
Characterizing nanomaterials

Technical Note

Resolution of a 3D X-ray Microscope

Defining Meaningful Resolution Parameters

Author: Carl Zeiss X-ray Microscopy, Inc.

Date: January 2019

Although the definition of resolution may seem straightforward, several non-equivalent terms are used to
represent the resolution capabilities of 3D tomographic instruments. ZEISS specifies spatial resolution, the
most meaningful measurement of a microscope’s performance. This Tech Note explains how to objectively
evaluate the resolution performance of 3D X-ray microscopes (XRM) and other computed tomography (CT)
instruments relevant to the user’s research and production goals.

Micro-CT and nano-CT technology have made considerable
strides in resolution during the past decade. The recent
introduction of 3D X-ray microscopy (XRM) brings yet
another advancement at even higher resolution.
When evaluating these high resolution systems it is helpful
to understand the numerous terms used in the industry to
describe resolution that, if not clearly understood, can
confuse comparisons between systems. These terms include:
• Spatial resolution • Spot size
• Voxel size • Detail detectability
• Nominal resolution
Each of these terms conveys vastly different representations
of a system’s performance. While spatial resolution provides
a direct measurement of the system’s complete imaging
capability, metrics such as voxel, nominal resolution, and
spot size are isolated contributors to resolution that,
by themselves, do not describe the performance of the
system as a whole.
Meaningful resolution measurements should reflect
total system performance, be based on standards that
enable unbiased comparisons, and be considered at
multipleworking distances that reflect the range of
intended applications and sample sizes. In order to
make an objective comparison of systems, a clear
understanding of each term is required.
Spatial Resolution: The Most Meaningful
Measurement
Spatial resolution refers to the minimum separation
at which a feature pair can be resolved by an imaging
system. It is typically measured by imaging a standardized
resolution target (Figure 1) with progressively smaller
line-space pairs. As the feature pairs become spaced
closer than a system’s resolution capability, they cannot
be distinguished (resolved) from one another and
increasingly appear as one single feature (Figure 2).

Figure 1 A standard spatial Figure 2 Spatial Resolution as a Function of Feature Separation. As the same pair of features becomes separated by
resolution taraget. spacing smaller than the resolution of a system, it becomes indistinguishable as a pair by the imaging system.

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Characterizing nanomaterials
Historically, spatial resolution came from attempts to
describe a telescope’s resolution by the minimum separa-
tion of stars that could be resolved. It still serves as the
standard scientific measurement of resolution for most
imaging systems, used for everything from medical CTs
to common optical microscopes.
Spatial resolution is the most meaningful and compre-
hensive metric because it measures the output of the
system (an image) and accounts for all imaging system
characteristics, including X-ray source spot size; detector
resolution; vibrational, electrical and thermal stability;
magnification geometry; and imaging conditions. This is
the metric that ZEISS uses to specify resolution of the
Xradia Versa and Xradia Ultra X-ray microscopes as
well as Xradia Context microCT.
Voxel: A Calculation of Pixelation
Voxel (3D) or pixel (2D) size is a geometric calculation
referring to a cross-sectional area in the sample that is
imaged onto a single detector pixel. This calculation accounts
for only the detector pixel size and system geometry, and
does not account for source blur; source and system stability;
imaging artifacts; and other key aspects of a system’s actual
output that contribute to or detract from image quality.
The fallacy of relying on a geometric term such as this can
be seen in Figure 3 in side-by-side images of a carbon fiber
composite at a voxel size of 1.0 µm. Although both images
were taken at the same voxel on different systems, only the
right-hand image can resolve the carbon fibers.
Pd
Dds (Detector
Pixel Size)
Dss
(V)
Detector Pixel
Sample
Source Rotation
Location
Technical Note
50 µm 50 µm
Figure 3 Voxel Size versus Spatial Resolution. Although the same carbon fiber
composite sample was imaged at the same voxel size of 1.0 µm, the resulting
image quality differs greatly. [Left: non-ZEISS commercially available CT system
at 1.0 µm. Right: ZEISS Xradia Versa at 1.0 µm
Minimum Voxel: Theoretical versus
Practical Calculation
Minimum voxel size is calculated assuming an extremely
small sample placed as close to the source and as far from the
detector as possible to deliver maximum geometric magni-
fication. However, operating at high geometric magnification
causes problems that can limit true resolution, such as source
blur and motion blur that can occur due to heating if the
sample is placed too close to the source.
Thus the minimum voxel size is often meaningless in depicting
the system’s actual resolution performance. In fact, a mini-
mum voxel may produce lower quality and worse resolution
than images taken at a larger voxel size.
It should be noted that while minimum voxel can be mis-
leading, the effective voxel can be relevant. When voxel size
becomes approximately equivalent to all other resolution
limiters, it is referred to as the effective voxel. This is
discussed further in the spot size section.
Nominal Resolution: “In Name Only”
Nominal means ideal or theoretical and is used to emphasize
the difference between theory (system design) and reality
(system performance). Nominal resolution is a theoretical
specification, and does not provide evidence of a system’s
true performance. In the practice of microCT the term is often
used to refer to the minimum voxel size. A common oversight
is to assume nominal resolution is equivalent to the spatial
resolution of a system; rather it is a purely geometric term
that becomes meaningless when other system resolution
factors, such as stability or source blur, are dominant.
8
Characterizing nanomaterials

Technical Note

Spot Size: Only Relevant for Geometric

Magnification-Based Systems
Spot size is the measured diameter (full width at half Blurring
maximum, FWHM) of the X-ray beam at the source. Source
Blurring
It is one of many factors that define a system’s ultimate
spatial resolution, but its impact is highly dependent
on a system’s optical and imaging design.
Sample
The source spot size relates to resolution due to
spot-size-induced image blur, referred to as “spot blur”
or “penumbral blur,” which is proportional to both the Figure 4 Spot Blur at Varying Geometric Magnifications. Spot blur can be
significant at high geometric magnifications (left), but can be significantly
source spot size and geometric magnification. This spot reduced when operating at lower geometric magnifications (right).
blur can significantly limit resolution (as seen in Figure 4)
for geometric magnification-based systems. As a result,
ZEISS Xradia
conventional micro-CT and nano-CT manufacturers have
Traditional uCT 600-series Versa
focused on developing the smallest system spot size.

The problem with this approach is that it has created a

Power / Throughput

race to the smallest spot size which has been badly plagued
by performance trade-offs. As a result, the smallest spot
sizes advertised on many micro-CT or nano-CT systems are
achievable only under very limited operating conditions;
typically at low voltage, low power, and often requiring
Spot size
physical reconfiguration of the source to a special filament.
Consequently these ‘high resolution modes’ apply to only
a narrow band of imaging applications and are a poor
indicator of overall system performance for the broad
spectrum of samples. Low Resolution High

Figure 5 Projection-based microCT and nanoCT systems suffer from a

This concept is depicted in Figure 5. Due to the above-
mentioned trade-offs between spot size, resolution, and
imaging parameters, users of traditional CT systems are
forced to chose between high throughput (large spot size)
or high resolution (small spot size).
However, not all system architectures are equally dependent
upon spot size. Two-stage magnification (geometric + optical)
designs employed by ZEISS X-ray microscopes reduce depen-
dence upon geometric magnification and minimize spot blur,
which removes the conventional limitations of spot size
on resolution.
resolution-throughput tradeoff due to spot blur. X-ray microscopy systems
do not suffer this tradeoff because they are much less susceptible to spot blur.
This is again depicted in Figure 5. With the Xradia Versa
600-series, increased power is provided without compro-
mising on other aspects of performance. This means that
users can now experience the same high resolution and
image quality for which the Versa series is well-regarded,
now with more photons and higher throughput.
Thus, while spot size can limit resolution for geometric
magnification-based systems including conventional
microCT/nanoCT, it should not be used as a comparative
metric when the comparison includes systems such as
X-ray microscopes that use two stages of magnification
and are not solely dependent on geometric magnification.

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Characterizing nanomaterials
Detail Detectability: Non-Standard Term
Also known as “feature recognition,” the term “detail
detectability” is used in various ways among the imaging
community. Detail detectability, depending on how it is
defined, can be used to characterize system performance.
However, due to dissimilar definitions within the CT
community and the lack of a standardized reference
target, it generally cannot provide the complete basis
for evaluation.
This term has been used confusingly in different contexts
Resolution (µm)
to refer to minimum voxel, to half the source spot size,
or to the smallest high-Z feature that an observer would
interpret as detected. Defining detail detectability as the
smallest high-Z feature that can be detected is the most
conventional definition, but there is not a common
method of determination.
Such quantification of detail detectability is problematic
because it is influenced by:
• Observer bias: an observer may visualize a slight
grayscale shift and subjectively claim the particle is
“detected” but interpretation may vary by observer.
• Feature construction: there is no standardardized
reference target for micro-CT, which can lead to
dissimilar detail detectability numbers that are reliant
on variable measurement techniques. Such techniques
may include creating a feature from highly X-ray
absorbing materials with high aspect ratios
(small only in one of the three XYZ dimensions),
which then enables detail detectability claims equal
to the smallest feature dimension. In other words,
because no standard measurement technique exists,
relying on detail detectability as a measurement of
resolution in an evaluation may lead to erroneous
conclusions.
Even if a standardized test method is constructed in the
future, detail detectability is still a limited measurement
because of its subjective nature.
Technical Note
Figure 6 Detail detectability measures the presence of something without the
requirement to resolve that object. When is the feature, the letter R, detected
versus resolved in these images? Detection occurs in the second or third image
from the left, well below where the feature is resolved.
Resolution at a Distance (RaaD™)
0
2 Xradia Versa XRM
4
6
8
Geometric Mag-based microCT’s
10
12
14
0 10 20 30 40 50
Working Distance (mm) – Similar to sample radius
Source to center of sample rotation
Figure 7 ZEISS Xradia Versa deliver resolution at a distance, or RaaD™,
to maintain the highest resolution at a wide range of working distances.
Using RaaD, the resolution achieved by Xradia Versa, which are maintained
at around ~1 μm, remains high while the resolution of micro-CT and nano-CT
rapidly fall off to coarse representations. ZEISS delivers the highest resolution
for in situ studies with flexibility for a wide range of sample sizes, conditions,
and distances.
The Distance Factor
When evaluating the performance of an X-ray imaging
instrument or comparing capabilities of several different
instruments, measurements of spatial resolution should not
just be assessed under best case scenarios for a system.
Resolution can degrade as working distance increases so
it should be evaluated across realistic imaging conditions
that include larger working distances for a wide range of
samples sizes relevant to the user’s intended applications.
For example, if one wishes to image a range of sample
sizes in 10 to 100 mm diameters, or conduct in situ
experiments to observe samples under specific conditions
such as temperature changes or compression and tension,
then spatial resolution at working distances appropriate
to those dimensions will be most relevant. By obtaining
spatial resolution measurements at multiple working
distances, a proper evaluation can be made for the
resolution achievable across the range of samples the
user cares about most.
10
Characterizing nanomaterials

Technical Note

Summary: Comparing and Selecting a 3D X-ray

Imaging Instrument
• Spatial resolution provides the most meaningful
method of evaluating an instrument’s performance,
and thus should be used to compare systems objectively.
• While minimum voxel, nominal resolution, spot size, and
detail detectability each can provide some information
about the system’s ability to resolve an image, each is
incomplete on its own and can misrepresent the true
imaging performance of the system.
• Spatial resolution should be assessed at working distances
relevant to the user’s intended applications.
• To choose the correct system, users must apply consistent
definitions while evaluating imaging solutions.
• ZEISS XRM deliver the highest spatial resolution across
the widest range of working distances.

Cover Image: Sample courtesy of Kavan Hazeli, Mechanical and Aerospace Engineering, The University of Alabama, Huntsville.

11
Characterizing nanomaterials

Technical Note

In Situ Observation of Mechanical Testing

at the Nanoscale
 Characterizing nanomaterials

Technical Note

In Situ Observation of Mechanical Testing

at the Nanoscale
Author: Carl Zeiss X-ray Microscopy
Pleasanton, California

Date: March 2015

X-ray microscopy (XRM) enables nondestructive 3D investigation of a variety of samples across multiple length scales.
By adapting advanced X-ray optics from synchrotron developments, 3D X-ray tomography with resolution down to
50 nm is now available in the laboratory. As a nondestructive technique, X-ray tomography uniquely enables 4D studies
by observing 3D structure at multiple points in time and under varying realistic conditions. These conditions can
include a variety of external stimuli, including mechanical loads. In this Technical Note we introduce the concept of
in situ mechanical testing integrated in nanoscale XRM. This approach is complementary to established ex situ or
in situ testing methods in SEM or TEM, but covers a unique 3D length scale, offering new opportunities to connect
small scale evolution processes with those observed in micron scale XRM and bulk material testing. This connection
of micro and macro deformation behavior, with direct 3D visualization, has promising applications covering a variety
of materials from metals to biomaterials to thin coatings. Xradia Ultra Load Stage, a new nanomechanical test stage,
is now available for integration into ZEISS Xradia Ultra, the only nanoscale XRM available for the laboratory.

Nanomechanical testing seeks to answer the question:
How does a material yield, deform, and fail on small scales?
By combining with in situ 3D observation, we now seek to
answer the next question: how do these mechanisms vary locally
within the sample, and what role does the microstructure play
in determining the bulk properties and behavior?
This broad concept can be further refined with several more
targeted questions:
• How is the behavior of individual micro- and nano-features
such as struts, films, “walls”, particles, etc. different from
the collective bulk? How does this limit the interpretation
of material properties from a purely geometric perspective
based on simple morphological properties like porosity,
surface area, etc.?
• What effects do these features have on the local deformation
of a material? For example, do some locations display brittle
behavior and others ductile? Do these events occur with
sufficient frequency or prevalence that the effects cascade
up to larger length scales?
• In terms of engineering a material with desirable properties
(modulus, hardness, toughness, etc.) are some defects and
features acceptable and others not?
Investigating Under Load
To investigate the deformation and possible structural failure
under various loading conditions, a nanomechanical testing
rig has been designed that can be easily configured by the
user to enable three different operating modes—compression,
tension,and nanoindentation. Xradia Ultra Load Stage operates
on samples of sizes typical of the nanoscale X-ray microscope,
on the order of a 10-100 micron diameter cross section.
This length scale simultaneously satisfies two experimental
constraints by:
1) providing optimal X-ray attenuation characteristics for a
wide variety of sample types, and
2) effectively connecting the length scales of material
deformation spanning resolutions from tens of nanometers
up to sample sizes approaching bulk material response.

13
 Characterizing nanomaterials

Technical Note

The Nanomechanical Testing Gap

The Ultra in situ load stage explores a new critical length scale 3D Internal / microCT Submicron
Bulk XRM Nanoscale
of materials characterization, observing the types of internal XRM
features, such as nanoscale cracks and voids, that initiate
as well as accentuate material deformation and failure, and Thin /
Transparent TEM
subsequently connecting these features with properties that
LM
are observed on the macroscale.

2D Surface SEM
This new capability satisfies a void in the current availability of
mechanical testing and imaging approaches. On the spectrum
of force and length scales, Xradia Ultra Load Stage is positioned 5 µm 500 nm 50 nm 5 nm

between, on the one hand, that of macro scale to microtomog- Resolution

raphy testing1, and on the other, established nanomechanical
testing such as in situ SEM, TEM2,3 (Figure 1), or standalone
nanoindenters. Of the currently existing in situ techniques,
micron scale X-ray tomography provides spatial resolution down
to the single-micron or submicron scale and can accommodate
samples on the millimeter or larger scale subjected to kilonewton
loads. On the small end of thespectrum, electron microscopy
enables in situ mechanical testing on the nanometer scale.
Despite providing excellent spatial resolution, the electron
techniques suffer from some inherent limitations. SEM is capable
of nondestructive surface imaging only, and TEM can provide
3D data but only on very thin samples, typically less than one
micron, where surface effects dominate the deformation
behavior and strongly influence the results. For both EM
methods, there is a lack of understanding of internal micro
and nanoscale deformation within a sample which is sufficiently
large to minimize surface effects. Conversely, the larger sample
sizes typical of the Xradia Ultra microscope (10-100 microns)
approach, in many instances, bulk material behavior, providing
a new length scale of investigation as well as complementary
information to connect length scales.
Figure 1. Approximate imaging resolution for common in situ mechanical testing
methods, categorized by sample thickness and transparency. The in situ Xradia
Ultra Load Stage uniquely fills a gap between single nanometer resolution
SEM/TEM methods that are restricted to surface imaging or extremely thin
samples, and micron/submicron-scale tomography.
Modes of Operation
Next, we focus on the following three modes: compression,
tension and indentation. In the implementation of these three
modes in nanoscale XRM (ZEISS Xradia Ultra), the sample is
mounted between two anvils, one of which is stationary and
one of which can be moved in a controlled manner. A sensor
measures the force on the sample as a function of anvil
displacement.
• Compression: the sample is “pinched” between the two
anvils that move towards each other. In this mode, structural
deformation under uniaxial compressive load is studied.
• Tension: the two anvils move away from each other, pulling
on the sample. In this mode, structural deformation under
uniaxial tensile load is studied.

• Indentation: a sharp nanoindenter mounted to one of the

anvils is pushed into the sample to initiate cracks, delaminate
coatings etc. Common types of indenters include cone,
cube corner, or wedge.

14
 Characterizing nanomaterials

Technical Note

Compression
In situ nanomechanical compression combined with X-ray to- Compression
mography can be used to observe the deformation of materials
under compressive load. This type of mechanical test has been
Top Anvil Top Anvil
commonly used in ex situ applications to study the behavior of
samples such as metal pillars and porous foams, but a direct 3D
image of the deformation process on this length scale has not X-ray X-ray
been previously available. This has left a number of unanswered Beam Path Beam Path
questions in terms of the relevant local mechanisms. Do micro
Sample Sample
and nanoscale features undergo uniaxial compression in a similar
manner to the bulk sample, or are there local buckling/yielding
events? Likely there is varying behavior especially for small, high
Bottom Anvil Bottom Anvil
aspect ratio features such as struts or pore walls. Pores of differ-
ent sizes may also yield at different rates. Furthermore, if there
are anisotropic structures what is the relationship between load-
Sample Rotation Sample Rotation
ing direction, deformation direction, and feature orientation? for Tomography for Tomography

These types of structures could have implications for the lateral

expansion of the sample orthogonally to the loading direction.
And finally, how do these microstructures affect relaxation after
load? It is now possible to observe if certain features relax
elastically, while others remain permanently deformed.

Tension
Tension
Tensile testing can reveal an additional wealth of information
about a sample’s deformation and failure mechanisms, including
critical properties like elastic modulus and tensile yield strength.
Top Anvil Top Anvil
While these properties can be traced back to atomistic level
deformation events in the form of dislocations, at what level
do these processes combine in aX-ray
fashion that dictates bulk X-ray
Beam Pathcan uniquely help address Beam Path
behavior? Xradia Ultra Load Stage
this question by operating at an intermediate length between
Sample Sample
atomistic information and bulk properties; at this scale nano/
microstructural features can constitute critical locations where
isolated deformation events likely
Bottom begin to occur on a length
Anvil Bottom Anvil
scale capable of propagating up to macro-level response.
In a tensile scenario, these types of critical locations include
micro-voids, where increased loading can lead to growth and
Sample Rotation Sample Rotation
for Tomography
coalescence of neighboring voids, as well as possible crack for Tomography
initiation sites in high stress regions of surrounding material.
It can also include thin and anisotropic features such as struts
and fibers which can undergo local rupture and failure at differ-
ent rates and under different loading than larger domains. It is
also possible, particularly in the case of composite materials, to
consider regions of varying chemistry or grain structure, resulting

15
 Characterizing nanomaterials
possibly in effects such as locally ductile expansion in one region,
but locally brittle failure in a neighboring region. These detailed
microstructural effects could indeed have implications for a wide
variety of sample types including the life sciences (bone, natural
Top Anvil
fibers, animal setae, hair), geosciences (mechanics of rock and
ore), and materials science (metals, polymers, ceramics, MEMS).
X-ray
Indentation Beam Path
Nanoindentation offers the opportunity to study very isolated
Sample
deformation and failure events surrounding the indentation site.
This type of test has a number of distinct advantages, including
the study of samples with small linear dimensions, such as coat-
Bottom Anvil
ings and layered structures. In addition, the capability to apply
the load in a carefully isolated manner enables the user to locally
select the position where failure should initiate. For example, in
Sample Rotation
a composite material consisting of a matrix and distributed inclu-
for Tomography
sion particles, indentation provides the researcher with the abili-
ty to initiate deformation of the matrix either in close proximity
or extended distance from an inclusion. In a general sense, the
indentation can be targeted near particular features of interest,
like voids, micro-channels, or interfaces. The exact location in
which force is applied often has great impact on such phenome-
na as crack initiation, growth, and coalescence.
By combining the indentation test with the 3D imaging capability
of nanoscale X-ray microscopy, it is possible to observe these
processes as they occur not only on the surface but also within
the sample interior. For thin films and coatings, this can provide
insight into localized delamination processes that occur at
sub-surface interfaces. For a variety of additional sample types
such as metals, concrete building materials, bone, and teeth,
it reveals the impacts that internal heterogeneities and defects
near the indentation site have on the deformation and failure
of the overall sample.
Practical Considerations
Sample sizes for the described methodology are on the order
of tens to hundreds of microns, and based on the Young’s mod-
ulus for elastic deformation over a range of sample types, such
sizes typically require forces in the millinewton to single Newton
range to produce deformation on the micron scale. To track this
behavior quantitatively during loading, load-displacement curves
are acquired to relate the applied force to the deformation or
damage events directly observed from the imaging.
Technical Note
Indentation
Top Anvil Top Anvil
X-ray X-ray
Beam Path Beam Path
Sample Sample
Bottom Anvil Bottom Anvil
Sample Rotation Sample Rotation
for Tomography for Tomography
As another consideration, data acquisition times in lab based
nanoscale XRM are on the order of hours for 3D tomographic
datasets. Therefore, real time observation of dynamic processes
is not practical. Instead, nanoscale XRM lends itself to so-called
“interrupted 4D” experiments, where 3D datasets are acquired
at multiple, discrete steps when the loading condition is held
static. The in situ configuration is still critical for this type of
measurement as the load must be maintained during imaging
to properly represent deformation processes such as the opening
of crack tips. To advance scientific understanding during dynamic
loading between tomography acquisitions, the sample can be
monitored in 2D radiography mode on the time scale of seconds
to minutes.
Lastly, while X-ray tomography ideally is performed over a
rotation range of at least +/- 90 degrees, nanomechanical testing
requires a structurally rigid connection of the two anvils between
which the sample is mounted. The lack of mechanically strong
but X-ray transparent materials in the X-ray energy range of
interest (5.4 or 8 keV in currently available instruments) requires
a narrow but strong support post. This limits the angular range
for tomographic data acquisition to about +/- 70 degrees.
16
 Characterizing nanomaterials

Technical Note

Demonstration
The concept of in situ nanomechanical testing in XRM is demon-
strated here on two types of materials intended to indicate the
possible range of applications. A compression experiment was
performed on a porous elastomer as shown in Figure 2. The
sample is mounted on the bottom anvil (not visible) and occupies
most of the field of view, while the stationary top anvil is also
visible at the top of the field of view. The three panels display
the results of three separate tomography scans conducted be-
fore, during, and after a compressive loading process. Panel
(a) depicts the original, uncompressed sample. In panel (b), the
sample has been moved vertically upward, putting the sample
in contact with the top anvil causing compression. The loading
was maintained in this state during the collection of the tomog-
raphy. After tomography, the sample was moved back down to
the original position to remove the compressive force. A third
tomography, depicted in panel (c), reveals the final deformation
of the sample after removal of the applied load, suggesting
permanent changes of the microstructure.
In a second experiment, nanoindentation was performed on a
sample of dentin. Dentin is a natural microstructured composite
material found in teeth, serving functions of transport, cell
communication, and structural reinforcement. The material
displays excellent mechanical and fracture toughness, making
it a prime candidate for biomimetic engineering. In this work,
in situ X-ray microscopy was performed with Zernike phase
contrast to observe both the sample and the diamond indenter
tip. Evidence of the internal tubular dentin structure, which
is hypothesized to be responsible for many of the material’s
impressive properties, is also visible from the surface renderings.
Two data sets were collected, first at light load and second after
partial fracture initiated by cracking around the indenter tip, as
shown in Figure 3 below. Investigation of the internal structure
can reveal where these cracks initiate and propagate relative
to the tubules, helping to explain why bulk dentin displays
such exceptional mechanical toughness.

a b c

10 µm

Figure 2. Compressive loading of a porous elastomer. (a) Uncompressed, (b) Compressed, (c) Decompressed

a b

10 µm

Figure 3. Nanoindentation of dentin. (a) 3D rendering of dentin sample before cracking. The indenter tip is visible above the sample.
(b) 3D rendering of the same sample after the indenter tip has been driven into the sample and a piece of dentin has fractured off.
(c) Force-displacement curve. The fracture event is clearly visible. Courtesy The University of Manchester.

17
 Characterizing nanomaterials

Technical Note

Outlook
In situ mechanical testing in a nanoscale X-ray microscope
applies to a wide range of interests, covering both engineered
and natural materials. Many such materials display hierarchical
structures, with the behavior at the bulk scale intimately linked
to the structure and properties of multiple smaller scales.
Nanoscale XRM, sitting in the unique nanometer to micron
nondestructive 3D imaging regime, has the potential to help link
observations across this vast range of scales, and complement
existing methods including in situ SEM, TEM, and micron scale
XRM. Specifically, Xradia Ultra Load Stage, capable of compres-
sion, tension, and indentation loading, offers new capabilities
to observe internal processes such as elastic and plastic deforma-
tion, crack initiation and propagation, and surface delamination
down to the 50 nm scale. While not an inclusive list, this is
envisioned to have strong applications in such fields as high
strength alloys, coatings, fiber composites, biomaterials,
building materials, and engineered foams.

References:
[1] BM Patterson, et al., “Measure of Morphological and Performance Properties in Polymeric Silicone Foams by X-ray Tomography,” J Mater Sci 48 (2013):
1986-1996. DOI: 10.1007/s10853-012-6965-2
[2] W Kang, et al., “In-Situ Uniaxial Mechanical Testing of Small Scale Materials--A Review,” Nanoscience and Nanotechnology Letters 2 (2010): 282-287.
DOI: 10.1166/nnl.2010.1107
[3] MA Haque and MTA Saif, “In-situ Tensile Testing of Nano-scale Specimens in SEM and TEM,” Experimental Mechanics 42 (2002): 123-128.
DOI: 10. 1007/BF02411059
7

18
Characterizing nanomaterials

Application Note

50 µm

Characterization of Powdered Bulk Materials

Using Light and Scanning Electron Microscopy
Characterizing nanomaterials

Application Note

Characterization of Powdered Bulk Materials

Using Light and Scanning Electron Microscopy
Authors: Thomas Rieger, Tim Schubert, Dr. Timo Bernthaler,
Prof. Dr. Gerhard Schneider
Aalen University of Applied Sciences – Technology and Industry

Date: November 2018

The bulk density and flow behavior are important parameters for characterizing powders used in powder-
technology-based production processes such as the additive laser powder bed fusion process because these
parameters essentially determine the workability of the powder across a wide range of production processes.
[Fra16] Good flow behavior is of particular importance for powder-bed-based additive production processes for
handling the powder and applying the powder layers, while a high bulk density is extremely important for ma-
nufacturing non-porous components. [Kru15], [Tan17] Light and scanning electron microscopy combined with
the image analysis tool in the ZEISS ZEN 2 core software make it possible to efficiently characterize powders
and determine their quality.

Introduction
The bulk density and flow behavior of a powder are essenti-
ally determined by the powder morphology, topography, size
distribution and internal particle structure. While spherical
particles possess positive flow properties, irregularly or
needle-shaped particles have a less favorable flow behavior
due to possible interlocking processes between the particles.
Nevertheless, closer inspection of a spherical particle may
reveal a rough surface structure, which causes high friction
Experiment
Two granulated tungsten carbide cobalt powders with iden-
tical composition but very different bulk densities are avail-
able for testing. The bulk density of the two powders was
determined in accordance with DIN EN ISO 3923:2010 and
the flowability in accordance with DIN EN ISO 13517:2013
and shown in Table 1.
Bulk density [g/cm³] Flowability [50 g/s]

forces that greatly impact the movement of the powder Powder 1 – WC-Co (88-12) 4.51 29.0
Powder 2 – WC-Co (88-12) 5.89 28.5
when it flows or compacts. [Sch14] Dry powders with a high
proportion of fine particles also develop high adhesive forces Table 1 Result of the bulk density testing and the flowability testing

between the individual particles in the form of van der Waals

interactions that also restrict the flowability of the powder.
[Mol00] Furthermore, the internal particle structure has a
decisive influence on the bulk density and the stability of a
powder. For example, internal porosity reduces the bulk den-
sity as well as the particle stability. [Sch14] When testing the
suitability of a powder or comparing different powders to
one another, critical factors such as the morphology, topo-
graphy, size distribution, and internal structure must be exa-
mined to determine whether they are appropriate for subse-
quent production processes.
Powders were inspected using a ZEISS Axio Zoom.V16 stereo
zoom microscope at 80 times magnification in transmitted
light to investigate the difference in the bulk density.
A few grams of the particular powder were distributed ho-
mogeneously in a petri dish and then large-area tiled images
were created using a software function in ZEISS ZEN 2 core
to ensure a minimum number of 6,000 particles for each
analysis. An area of 3×3 individual images was defined. After
defining the classes and the measurement frame, the auto-
matic segmentation was carried out using gray level
thresholding with the help of the integrated image analysis
tool. Incorrect segmentation could be interactively corrected
before the analysis. The equivalent diameter, which corres-
ponds to the diameter of an ideal round particle that is

20
Characterizing nanomaterials

Application Note

equal in area, to determine the particle size as well as the definitive measurement characteristics. Figure 1 shows
the Feret ratio, which corresponds to the ratio of the smal- a section of a powder in transmitted light, left before seg-
lest to the largest Feret diameter and thus provides informa- mentation, right after automatic segmentation using the
tion about the sphericity of the particle, were measured as image analysis tool provided in ZEISS ZEN 2 core.

Figure 1 Powders for characterization using the ZEISS ZEN 2 core image analysis tool; left: not segmented, LM, 80×; right: segmented, LM, 80×

The particle size distribution and the Feret ratio distribution

could be determined using the many particles investigated
(> 6,000 objects) and depicted graphically; see Figure 2.

Figure 2 Left: particle size distribution; right: Feret ratio distribution comparing WC-Co powders 1 and 2

Considering the particle size distribution and the Feret ratio
distribution, the difference in the bulk densities must be at-
tributed to the internal structure of the particles because the
curves differed only very slightly. For this reason, the particles
were examined further using the ZEISS Crossbeam 550
focused ion beam scanning electron microscope (FIB SEM).
The powders were scattered onto a sample holder
smeared with graphite paste and sputter-coated with a
gold-palladium layer to ensure good electron conductivity.
Figure 3 shows an overview image, while Figure 4 shows
a detailed image with a comparison of the two WC-Co
powders.

21
Characterizing nanomaterials

Application Note

Figure 3 Overview image of WC-Co particles; left: powder 1, SEM, BSE signal, 250×; right: overview image of powder 2, SEM, BSE signal, 250×

Figure 4 Detailed image of the WC-Co particle topography left: powder 1, SEM, BSE signal, 2,500×; right: powder 2, SEM, BSE signal, 2,500×

It can be clearly seen in both the overview images and the
detailed images that powder 1 has an outwardly open poro-
sity, while powder 2 has an outwardly closed granulate sur-
face. Powder 2 also has a visibly finer surface structure that
can be attributed to a smaller WC particle size. To characteri-
ze the internal particle structure, a FIB section was placed
through the middle of a randomly selected particle with the
ZEISS Crossbeam 550 for each type of powder. The images
are shown in Figure 5 for comparison. Examining the images
suggests that powder 1 already has a considerably higher
fraction of internal porosity than powder 2.

22
Characterizing nanomaterials

Application Note

Figure 5 FIB section through particle left: powder 1 with higher internal porosity, FIB, 2,500×; right: powder 2 with lower internal porosity, FIB, 2,500×

For the quantitative determination of the porosity fraction,
the samples were prepared by embedding a few grams of
the powder in epoxy resin and the samples were then prepa-
red with various grinding and polishing steps using the
Struers Tegramin-30 system. The particles were cut during
the grinding and polishing steps, which provided a view of
their internal structure. To ensure the electrical conductivity
of the metallographic polished samples, a sputter-coating
process was carried out to deposit a gold-palladium layer
and finally the samples were viewed using a ZEISS Sigma
300 VP field emission scanning electron microscope (FE-SEM).
The pixel size of the images is 50 nm in order to cover the
smallest pore to be detected with a minimum number of
10 pixels. The additional analysis was also carried out using
the image analysis tool in the ZEISS ZEN 2 core software. A
hierarchical segmentation of the two gray level thresholds
was used, which initially enables the particles (red) and then
the internal pores (blue) to be segmented using an additional
gray level threshold. Figure 6 shows the classes which were
applied for the image analysis.

Figure 6 ZEISS ZEN 2 core image analysis tool; hierarchical arrangement of the classes

23
Characterizing nanomaterials
The measurement characteristics to be determined were
defined as the area of the individual particles and the area
of the internal pores present. Following allocation of the
pores to the particular particles, the fraction of internal
porosity could be determined and the mean porosity of
Powder 1
SEM image
Segmented using image analysis tool
Figure 7 Quantitative determination of the porosity fraction; top left: powder 1, SEM, BSE signal, pixel size 50 nm; top right: powder 2, SEM,BSE signal, pixel
size 50 nm; bottom left: powder 1 segmented, bottom right: powder 2 segmented
Application Note
the 20 particles of each powder examined could thus be
calculated. Figure 7 shows a section of the SEM image and
the associated segmented image using the image analysis
tool of the ZEISS ZEN 2 core software. The higher porosity
fraction of powder 1 is clearly apparent.
Powder 2
24
Characterizing nanomaterials

Application Note

Mean porosity [%]
Powder 1 – WC-Co (88-12) 34.7
Powder 2 – WC-Co (88-12) 4.8
Table 2 Result of the porosity analysis of the different WC-Co powders
The fact that the bulk density of powder 1 is > 20 % lower
can thus be explained by a mean porosity that is approxi-
mately 30 % higher. The flowability of the two powders is
almost identical thanks to the spherical particle shape of the
two powders. The somewhat better flowability of powder 2
can be attributed to the finer surface topography and the
resultant lower friction between the particles.
Summary
The option for characterizing powders described here reveals
that a powder can be almost fully characterized using light
and scanning electron microscopy. The image analysis tool of
the ZEISS ZEN 2 core software also allows quantitative hier-
archical evaluation of images from light microscopy and
scanning electron microscopy. Particles, pores and phases
can be automatically segmented, corrected and then analy-
zed using gray level thresholding. The automatic segmentati-
on function in particular enables simultaneous analysis of a
number of particles and thus provides rapid and statistically
reliable information about the quality of a powder.

References:
[Fra16] L. F. Francis, B. J. H. Stadler, C. C. Roberts: Materials processing, Elsevier (2016)
[Kru15] J.-P. Kruth, S. Dadbakhsh, B. Vrancken, K. Kempen, J. Vleugels, J. Van Humbeeck: Additive Manufacturing of Metals via Selective
Laser Melting, in Additive Manufacturing: Innovations, Advances, and Applications, Ed.: T. S. Sudarshan, et al., CRC Press (2015)
[Mol00] H. Mollet, A. Grubenmann: Formulation technology: Emulsions, suspensions, solid formats [Original in German: Formulierungstechnik:
Emulsionen, Suspensionen, Feste Formen], WILEY-VCH Verlag GmbH (2000)
[Sch14] D. Schulze: Powders and bulk materials [Original in German: Pulver und Schüttgüter], Springer Vieweg (2014)
[Tan17] Y. Tan, J. Zheng, W. Gao, S. Jiang, Y. Feng: The Effect of Powder Flowability in the Selective Laser Sintering Process, in Proceedings
of the 7th International Conference on Discrete Element Methods, Ed.: X. Li, et al., Springer Singapore; Imprint; Springer (2017)

25
Characterizing nanomaterials

Technology Note

Pt

Ag

Ni

Cu
2 µm 210 pA

200 nm

500 pA 1 nA

High Throughput Imaging with

ZEISS Crossbeam 550
Characterizing nanomaterials

Technology Note

High Throughput Imaging with

ZEISS Crossbeam 550
Author: Fabián Pérez-Willard
Carl Zeiss Microscopy GmbH,
Oberkochen, Germany

Date: March 2016

Introduction
Pt
Since more than 13 years ZEISS has been known as a
manufacturer of two-beam FIB-SEM systems – Crossbeam
Ag
instruments. During these years Crossbeam performance in
terms of image resolution and contrast, ease of use, work-
flow automation and throughput has improved continuously. Ni

This development culminates in today’s ZEISS Crossbeam 550. Cu

2 µm 210 pA

In this technology note, a simple experiment is presented to

illustrate the superb SEM imaging resolution of Crossbeam 550
when imaging simultaneously at low voltages and high
probe currents. Implications for different FIB-SEM applications 200 nm
are discussed, especially with regard to throughput.

Experiment
The sample used for this experiment consists – from top to 500 pA 1 nA
bottom – of a silver layer on a 1.5 µm-thick nickel seed layer
on a copper substrate [1]. A standard FIB cross-section of
the sample was produced (see top left image in figure 1).
Prior to FIB milling of the cross-section the sample surface
was protected with a platinum deposition layer by ion beam
induced deposition (IBID) using standard trimethyl (methylcy-
clopentadienyl) platinum (IV) as a precursor gas.

2 nA 4 nA
The subject of study of this note is the cross-sectioned
platinum deposit. The platinum deposit was imaged with the Figure 1 Images of a platinum deposition for different SEM probe currents
SEM at high magnification at two different accelerating in the range of 210 pA to 4 nA at 3 kV. (top left) Overview image of the cross-
section used for the experiment.
voltages, 1.5 kV and 3 kV, to reveal its internal structure.

27
Characterizing nanomaterials

Technology Note

Results
Figures 1 and 2 show the results for 3 kV and 1.5 kV,
respectively. For both imaging voltages, a granular structure
in the deposition is observed. This structure is well known
for FIB-SEM users. It consists of platinum nanoparticles
surrounded by a matrix of an amorphous carbon and gallium
mix. In the context of this note, it is used as a visual measure
210 pA 500 pA for the instrument’s SEM resolution. The platinum grains in
the images are well defined up to a SEM current of 2 nA.
With increasing current, they start to wash out at 4 nA, but
are still clearly resolved. For 1.5 kV and the highest current
200 nm of 8 nA the particles cannot be discerned.

Note that for 3 kV and 1.5 kV the SEM imaging current can
be increased from 210 pA to 4 nA roughly by a factor of 20
1 nA 2 nA
and – the platinum particles – the structure of interest – can
still be resolved.

The acquisition time for all the deposition images shown in

Figures 1 and 2 was either 5.4 s or 10.7 s.

Discussion
The results summarized in figures 1 and 2 demonstrate
Crossbeam 550 ability to maintain a very high resolution even
4 nA 8 nA
when imaging at very high SEM currents. This allows to boost
Figure 2 Images of a platinum deposition for different SEM probe currents throughput of the FIB-SEM instrument, as will be explained in
in the range of 210 pA to 8 nA at 1.5 kV (current increases from top left to
bottom right image).
the following for two common FIB-SEM applications:

A) Live imaging – For process control some applications rely

For each voltage the SEM probe current was varied in a
range from 210 pA to 8 nA. The images were acquired in the
coincident point of SEM and FIB beams, i.e. at a working
distance of 5 mm.
To exclude any contamination artifacts, every image shown
in this note was acquired from an area of the platinum
deposition, which had not been imaged or exposed to the
electron beam previously.
on live SEM imaging during FIB milling. Often live imaging
might not be strictly necessary, but it is still the preferred
mode of operation for ZEISS FIB-SEM instruments, since
it gives the operator the possibility to interact with the
instrument at any time and it has no drawbacks [2].
During live imaging a higher SEM probe current reduces
the impact of interferences caused by secondary electrons
produced by the scanning FIB beam. Thus, better SEM
imaging results can be achieved, or alternatively, the
milling process can be accelerated, because higher FIB
currents can be afforded without compromising the SEM
monitoring.

28
Characterizing nanomaterials

Technology Note

B) FIB-SEM tomography – As the need for 3D characterization Summary

of samples grows, FIB-SEM tomography is becoming
increasingly important: A volume of interest in the sample
is sliced by FIB and the resulting cross-sections imaged
with the SEM. Typically, material removal and imaging are
performed in a sequential manner. Experiments can run
from several hours up to several days. Meanwhile,
hundreds to thousands of SEM images are acquired.
In order to reduce the image acquisition time for a single
frame, and with it, for the complete 3D image stack, it is
important to choose the maximum possible SEM imaging
current, which still does not compromise the resolution
required to resolve the features of interest in the sample.
Cross-sections of platinum depositions were imaged with
ZEISS Crossbeam 550. The platinum nanoparticles present
in the depositions were used as a model system to show
the behavior of instrument resolution as a function of
current at low voltages. A very high imaging resolution
can be achieved for SEM beam currents up to several nA.
This makes Crossbeam 550 the tool of choice for high
throughput FIB-SEM work.

On the other hand, low voltages need to be employed to

ensure that the SEM information depth is comparable –
ideally less – than the thickness of the slices of material
removed by FIB. The information depth probed by the
SEM can be reduced to a few nanometers by detecting
only low energy loss backscattered electrons [3].

Recently, with the introduction of the advanced FIB-SEM

tomography solution Atlas 3D the sequential slicing
and imaging approach is no longer needed. Instead,
tomography data acquisition is done in live imaging mode
(see above), which allows to increase FIB-SEM tomography
throughput further. Researchers at NIH have reported
3 nm isotropic voxel resolution using this method [4].

References:
[1]
Sample courtesy of D. Willer, MPA Stuttgart, Germany.
[2
] All ZEISS FIB-SEM instruments have a non-immersion type objective lens. Therefore, live imaging is performed in normal imaging mode
as opposed to immersion type systems, where live imaging is only possible if the final lens is deactivated, which comes along with an
important loss in SEM resolution.
[3
] E. Schröder-Reiter et al., Journal of Structural Biology 165 (2009), 97 – 106; M. Cantoni and L. Holzer, Mrs Bulletin, 39 (2014), 354 – 360
[4]
K. Narayan et al., Journal of Structural Biology 185 (2014), 278 – 284

29
 Characterizing nanomaterials

Application Note

Exploring Laser-Assisted TEM Sample Preparation

with ZEISS Crossbeam laser
 Characterizing nanomaterials

Authors: Dr. Fabián Pérez-Willard, Tobias Volkenandt

Carl Zeiss Microscopy GmbH, Oberkochen, Germany
Date: 20 th of November 2020

This note explores the use of femtosecond (fs) laser ablation for site-specific TEM sample preparation in
a FIB-SEM. Different workflows are presented enabling TEM lamella preparation of regions of interest (ROI)
buried so deeply in the sample that they cannot be accessed directly by FIB. As a best-known method, the
so-called Cut-to-ROI workflow was developed and is explained in detail. General limitations when preparing
large TEM lamellas are discussed, as well as the new possibilities the LaserFIB opens for microstructural
characterization.

Introduction
Recently, LaserFIB instruments have attracted substantial
attention in the microscopy community. The term LaserFIB
denotes the combination of a FIB-SEM with an ultra-short
pulsed laser, typically a fs laser, within a single instrument.
The laser adds additional capabilities to the FIB-SEM,
expanding its already broad application space [1].
By means of fs laser ablation large volumes of material can
be removed from a sample very rapidly with in most cases
negligible sample damage. Thus, access can be gained to
regions of interest deeply buried in the sample for their
subsequent FIB-SEM analysis. Other applications of LaserFIBs
include the fabrication of micromechanical devices for materials
testing with dimensions of up to millimeters [2,3], the prepa-
ration of surfaces for EBSD analysis [4,5], and the production
of pillar-shaped samples for the synchrotron or X-ray microscope
[6].
In the LaserFIB setup implemented by ZEISS, with the product
name ZEISS Crossbeam laser, the laser work is done in a
dedicated chamber to prevent contamination of the main
FIB-SEM chamber by laser ablated debris [7].
In this paper we explore the possibilities ZEISS Crossbeam
laser offers for TEM sample preparation. We present different
preparation workflows and discuss possible modifications to
adapt to specific microstructural characterization challenges.
The workflows are well suited for the TEM cross section
preparation of deeply buried features of interest or the
preparation of multiple thinned windows distributed over
a large specimen area.
Experimental
All experiments were conducted on a ZEISS Crossbeam laser.
ZEISS Crossbeam laser is equipped with a diode pumped solid
state (DPSS) 515 nm (green) laser with laser pulses of less than
350 fs duration. Pulse repetition rates can be varied in the
range of 0.1 kHz to 1 MHz. The maximum output power is
10 W when operated at 1 MHz. With this laser, material removal
rates of more than 15 mio µm³/s in silicon can be reached,
representing at least three orders of magnitude improvement in
speed compared to any focused ion beam [1]. When defining the
laser milling strategy for a given layout, important parameters
are laser power (in percentage), writing speed (in mm/s), pulse
repetition rate and number of passes (hatches). The user can
input these for individual shapes or groups of shapes in a CAD
interface.
ZEISS Crossbeam laser features an Ion-sculptor gallium FIB
column with beam currents of 1 pA to 100 nA. For the lamella
lift-outs, an OP400 (Oxford Instruments plc) micromanipulator
and a gas injection system (GIS) were used.
The samples used were <100> silicon (for most of the
experiments) and a 99.9% pure copper foil (for the H-bar
experiment). These materials were chosen to illustrate the
different workflows because they are readily available,
homogeneous, and a good reference for comparison with
standard FIB work.

2
31
 Characterizing nanomaterials

Figure 1: SEM images of a laser-machined chunk. a) Top view with laser milling
shapes overlaid on one side of the chunk; blue for the rough milling and orange
for the polishing step. b) Magnified top view. c) Sample tilted 54° and rotated.

Chunking with the Laser

Figure 1 shows a laser prepared “macro-lamella”, also known as
a chunk, in silicon. The chunk was defined by first performing a
coarse milling step of two opposing boxes. One of these boxes
is schematically shown in Figure 1a) as a shaded blue rectangle.
In a second laser machining step – corresponding to the orange
shaded rectangle – both sides of the chunk were polished.

The coarse milling objects were nominally 400 µm wide. The

ones for the polishing step 350 µm. The position of the boxes
was chosen to yield a chunk thickness of around 10 µm as
measured at the top, see Figure 1b). After the laser polishing
step, the side walls of the chunk have a taper of (7±1)°.
Figure 2: SEM images of a 3 × 2 array of laser-machined chunks. a) Top view.
b) Sample tilted 54° and rotated. The FOV width is 2.3 mm for both images.
The preparation of the chunk by laser is analogous to standard
FIB preparation. The main difference is the scale. The dimensions
of the laser-prepared chunk are a factor of 10 to 20× larger than Thinning and Shaping by FIB
for a typical gallium FIB chunk. Still, the laser fabrication process The laser preparation was followed by gallium FIB machining
is very fast. For the chunk in Figure 1 the total laser machining to planarize and prepare the samples for lift-out. Different
time was 80 seconds. approaches were tested to explore their practicability.

In order to explore several different ways to use the laser
to assist TEM sample preparation, an array of 3 × 2 chunks
was prepared. This array is shown in Figure 2. The complete
array spans an area of 1.6 × 1.3 mm² and was patterned in
240 seconds.
Figure 3 shows a first example. One of the laser-prepared
chunks was thinned down to slightly less than 2 µm over an area
58 µm wide and more than 75 µm deep by gallium FIB milling.
This area appears bright when imaged with the SEM at 15 keV
landing energy, Figure 3a). The thinning of this lamella was
done in three steps performed at FIB currents of 65 nA, 30 nA,
and 7 nA. The sample was over-tilted and under-tilted when
machining the front and back side, respectively. The over-
and under-tilt angles were reduced from 4°, to 3°, to 2° with
decreasing milling current to ensure a homogeneous thickness
over the entire thinned window. The FIB thinning including
all three steps took 3 h in total. This is a time we consider
acceptable in light of the sample size and the fact that the
FIB milling can be fully automated.

3
32
 Characterizing nanomaterials

Alternatively, the 3 h time budget can be allocated just to the

cut-out step of the as-lasered sample, without any FIB thinning/
polishing. This case is shown in Figures 4c) and d). Again, a FIB
current of 65 nA was used. The sample is now 300 µm × 140 µm in
size, with a thickness of 10.5 µm at the top and getting thicker
towards the bottom due to the side wall taper of 7°.

Lift-Out
Despite their size, all three lamellas discussed in the previous
section could be easily lifted out of the bulk sample and
attached to a 3 mm copper grid compatible with standard TEM
sample holders following the standard in situ lift-out procedure
[8].

For visual comparison, Figure 5a) shows a standard-sized FIB

lamella (15 µm × 14 µm), and Figures 5b) to d) show the lamellas
from the previously described experiments, following the lift-out
step. The FOV is the same for all four images in this figure.

Figure 3: a) SEM images of a large FIB thinned lamella. a) Sample tilted 54° and
rotated. b) Top view of the thinned region.

For the next sample, the same amount of time was invested to
thin a 220 × 104 µm² area to 4.5 µm thickness including cut-
out, see Figures 4a) and b). This time all FIB milling operations
were conducted at 65 nA. The over/under-tilt angle was 3°.
Note that these settings would allow to produce a lamella of
the dimensions in Figure 3 in well below 1 h.

Figure 5: a) SEM image of a standard FIB lamella after in situ lift-out. b) to d)

SEM images of laser prepared lamellas of increasing size after in situ lift-out.
The lamella in d) was attached to the same grid as the lamella in a), which is
highlighted by the arrow.

Figure 4: SEM images of two large lamellas after cut-out. a) and b) show
different SEM perspectives – 0° and 45° stage tilt – of a 220 µm × 104 µm × 4.5 µm
lamella. c) and d) show a bigger lamella with dimensions of 300 µm × 140 µm.
This one is 10.5 µm thick at the top with a side wall taper of 7°.

4
33
 Characterizing nanomaterials

Limitations
The next step after the in situ lift-out is to thin the lamella
towards electron transparency. TEM analyses require very thin
samples with a thickness of 100 nm or less. It is a challenge to
obtain a uniformly thinned window of this thickness covering a
large area.

Depending on the sample material, FIB-thinned windows are

typically of the order of 10 µm × 10 µm or smaller. This is
because at some point during thinning the membrane starts
to bend. This renders further thinning impossible without
destroying the sample. The bending is due to intrinsic stress
release in the sample combined with its lower mechanical
stability. Thus, the material itself imposes limits to the achievable
size of an electron transparent window. This means that even
in large chunks only small windows can be thinned. However,
numerous small TEM windows can be prepared to cover a large
specimen area.

We see the main application of the LaserFIB for TEM sample

preparation, as providing three-dimensional (3D) site-specific
preparation of deeply buried structures of interest. The next
section describes how this can be accomplished using what we
have named the Cut-To-ROI workflow.

The Cut-To-ROI Workflow

The starting point or pre-requisite of the Cut-To-ROI workflow
is a laser-machined chunk. The chunk contains the ROI. Further,
the position of the ROI in the depth of the chunk (direction of
the bulk sample surface normal, Z) is known.

Figure 6: Laser processed lamella after FIB shaping to access ROIs 45 µm, 65,
The XYZ coordinates of a ROI can result from the sample lay- and 25 µm below the surface (dotted circles). b) The top surfaces after FIB milling
out itself (e.g. the known CAD layout of an electronics or semi- are flat and smooth. c) SEM image at 5 kV of a different lamella after thinning of
two ROIs.
conductor sample), from a previous characterization by a non-
destructive imaging technique (e.g. by X-ray microscopy) [9], or
any combination of both (e.g. lock-in thermography combined After trimming, the grid was flipped back to the upright posi-
with CAD chip layout). Any feature of interest that has been tion. Figure 6b) shows the top surface of one of the steps.
located in the sample can be targeted by the laser with an The surface is flat and smooth, which facilitates the subsequent
accuracy of about 2 µm [10]. preparation of the TEM-thin areas by FIB. Figure 6c) shows
another exemplary chunk after final thinning of two ROIs 25 and
The chunk can be trimmed by FIB as shown in Figure 6a). 45 µm below the surface.
Here, the grid was flipped by 90° after the lift-out, so that
FIB incidence was perpendicular to the chunk side face for
trimming. Three ROIs were assumed at Z = 45, 65 and 25 µm,
from left to right. The FIB was used to remove the top of the
chunk to form 40, 60, and 20 µm deep steps. This process took
13 min for the three steps at a FIB current of 30 nA.

5
34
 Characterizing nanomaterials

Figure 7 illustrates a variation of the Cut-To-ROI workflow.

In this case, the laser machined chunk was not lifted out of the
bulk but was trimmed in the Z-dimension directly by FIB. For this
purpose, the sample stage was at 0° tilt. The FIB trimming then
takes place at an angle of 36° to the bulk sample surface (XY
plane). In this case, our assumed ROI was at Z = 80 µm. The top
75 µm of the chunk were removed over the entire width of the
chunk (65 nA FIB current, 1h 36min FIB milling time) and then
two standard FIB in situ lift-out specimens were prepared.
For most applications, this alternative Cut-To-ROI workflow can
be sped up easily by preparing a thinner chunk, or even allowing
some material removal from the top of the chunk by the laser
beam tail.

Figure 8: H-bar preparation in a piece of copper foil by laser. Two large H-bars
were prepared. The one in the inset was machined in 34 seconds.

Summary
In this work different workflows were discussed combining
laser and FIB machining for the preparation of TEM specimens.
Figure 7: a) Laser-processed chunk. After laser cutting, the top 75 µm were ZEISS Crossbeam laser systems allow the 3D-site-specific
removed by FIB milling and two standard FIB lamellas were prepared. b) Detail of
the right-most lamella corresponding to the area in the white frame in a).
preparation of deeply buried regions of interest, or the
preparation of multiple TEM-suitable windows spread over
H-Bar Preparation by Laser a large sample area.
Another option is to combine H-bar [11] and Cut-To-ROI prepa-
rations. This technique can be applied e.g. on small pieces of
metal foils or wafers. These pieces are mounted on edge for laser
and FIB machining. This is shown in Figure 8. Two chunks were
patterned into a 300 µm thick copper foil using the laser. The
laser processing time for the larger chunk of 400 µm × 215 µm
was 34 seconds.

References
[1] F. Pérez-Willard et al., Expanding the application space of ion microscopy, submitted.
[2] M. J. Pfeifenberger et al., The use of femtosecond laser ablation as a novel tool for rapid micro-mechanical sample preparation,
Materials and Design 121 (2017), pp 109-118.
[3] M. J. Pfeifenberger et al., Electron Irradiation Effects on Strength and Ductility of Polymer Foils Studied by Femtosecond Laser-
Processed Micro-Tensile Specimens, Materials 12 (2019), pp. 1468-1483.
[4] M.P. Echlin et al., The TriBeam system: Femtosecond laser ablation in situ SEM, Mater. Charact. 100 (2015), pp 1-12.
[5] T. Schubert et al., Rapid Sample Preparation for EBSD-Analysis Enabled by the LaserFIB, ZEISS Application Note (2020), available
online.
[6] B. Tordoff et al., The LaserFIB: New Application Opportunities Combining a High-Performance FIB-SEM with Femtosecond Laser
Processing in a Second Integrated Chamber, Appl. Microsc. accepted.
[7] H. Doemer et al., Processing System, US8115180B2 (2012).
[8] L. A. Giannuzzi and F.A. Stevie, Introduction to Focused Ion Beams: Instrumentation, Theory, Techniques, and Practice,
New York, Springer (2005).
[9] M. Kaestner et al. Novel Workflow for High-Resolution Imaging of Structures in Advanced 3D and Fan-Out Packages, 2019
China Semiconductor Technology International Conference (CSTIC) (2019), pp. 1-3.
[10] ZEISS Crossbeam Product Specifications, available on request at www.zeiss.com/crossbeam
[11] J. Li, “Advanced Techniques in TEM Specimen Preparation”, in The Transmission Electron Microscope, Ed. Dr. K. Maaz,
IntechOpen (2012).

6
35
Characterizing nanomaterials

Technical Note

Diffraction Contrast Tomography

Unlocking Crystallographic Information
from Laboratory X-ray Microscopy
Characterizing nanomaterials

Technical Note

Diffraction Contrast Tomography

Unlocking Crystallographic Information
from Laboratory X-ray Microscopy

Author: Carl Zeiss X-ray Microscopy, Inc.

Date: January 2019

X-ray tomography has operated under two primary contrast mechanisms for some time: X-ray absorption
and phase contrast, which both rely on material density differences within the sample. However, single-phase
polycrystalline materials (e.g. steels, alloys, etc.) do not exhibit any significant contrast using absorption or
phase mechanisms. Synchrotron-based XRM has demonstrated results in this area for about a decade with
diffraction contrast tomography (DCT), which provides crystallographic/diffraction information from poly-
crystalline samples, non- destructively, in 3D. Now, advancing laboratory X-ray microscopy (XRM) one step
further, we describe here the capabilities of laboratory- based DCT on the ZEISS Xradia 620 Versa 3D X-ray
microscope, and the new research and characterization capabilities this enables.

Introduction
In the continued spirit of transferring synchrotron capabili-
ties to laboratory XRM systems, ZEISS in partnership with
Xnovo Technology, has implemented the ability to obtain
crystallographic (diffraction) contrast information on the
ZEISS Xradia 620 Versa [1,2].
Crystallographic imaging is commonly known from several
metallographic techniques, including light and electron
microscopy (EM) methods. In recent years, the introduction
of 2D and 3D electron back-scattering diffraction (EBSD)
techniques have made EM a routine tool for research and/
or development related to metallurgy, functional ceramics,
semiconductors, geology, etc. The ability to image the grain
structure and quantify the crystallographic orientation relation-
ships in such materials is instrumental for understanding and
optimizing material properties (mechanical, electrical, etc.)
and in situ processing conditions.
However, the destructive nature of 3D EBSD (via examination
of sequential slices) prevents one from directly evaluating the
microstructure evolution when subject to either mechanical,
thermal or other environmental conditions. Understanding
this evolution process on the same sample volume is key
to unlocking a more robust understanding of materials
performance, along with improved modeling capabilities,
and is a key driver of future materials research efforts.
In response to this constraint, synchrotron-based
crystallographic imaging, known as diffraction contrast
tomography (DCT) has emerged over the past decade [3,4].
Utilizing non-destructive X-rays, synchrotron users could
quantify grain orientation information in the native 3D
environment without physical sectioning. This has led
to the logical desire to study the evolution of grain
crystallography in situ or during interrupted “4D”
evolution experiments. However, limited regular access
to such synchrotron techniques has constrained the
ability to perform thorough, longitudinal studies on
materials evolution.

Cover Image LabDCT 3D imaging of grain orientations in aluminum alloy – IPF (inverse pole figure) colored.
Image Courtesy of: TU Denmark

37
Characterizing nanomaterials

Technical Note

ZEISS has taken this established synchrotron modality

and successfully transferred it to the ZEISS Xradia Versa family
of laboratory-based sub-micron XRM systems. LabDCT may
be efficiently coupled to in situ sample environments within
the microscope or subject to an extended time evolution
“4D” experiment (across days, weeks, months) – a unique
practical strength of laboratory-based XRM/DCT experiments [5].
Following an evolution experiment, the sample may be sent
to the electron microscope or focused ion beam (FIB-SEM)
for post-mortem complementary investigation of identified Figure 1 Schematic of the LabDCT setup. The sample is illuminated through
volumes of interest. This natural correlative workflow com- an aperture in front of the X-ray source. Both the sample absorption and
diffraction information is recorded with a high resolution detection system.
bining multiple lab-based methods on the same sample
Optionally, a beamstop maybe be added to the set-up in order to increase
volume remains an attractive future direction of microscopy the dynamic range of the diffraction detection.
to many researchers and is enabled by the ZEISS foundation
and breadth of imaging instrumentation.

LabDCT: How it Works

Here, we present a method to acquire, reconstruct and
analyze grain orientation and related information from
polycrystalline samples on a commercial laboratory
X-ray microscope (ZEISS Xradia 620 Versa) that utilizes a
synchrotron-style detection system.

A schematic representation of the LabDCT implementation

is shown in Figure 1. The X-ray beam is constrained through
a specialized aperture to illuminate the sample. Diffraction
information is collected with a tailor-made high resolution
detector that in addition allows simultaneous acquisition of
the sample’s absorption information. In order to increase the
sensitivity to the diffraction signal a beamstop may be added.
The data acquisition is performed in a symmetric geometry,
which enables improved diffraction signal strength and allows
handling of many and closely located diffraction spots.

Figure 2 Example 2D diffraction pattern from a Ti-alloy. The central area in the
With LabDCT a series of projections is taken from which projection is obscured by a beamstop. This missing data does not impact the
spatially resolved crystallographic information within the diffraction analysis and reconstruction.

sample is derived. Figure 2 illustrates a single such 2D

diffraction pattern as acquired with the LabDCT detector.
Just a few hundred projections are enough for a complete
LabDCT data set.

38
Characterizing nanomaterials

Technical Note

Validation and Application Examples

Polycrystalline Ti-alloy (Timet 21S) and Al-alloy samples A B

have been explored and are used here to demonstrate

the results of LabDCT. Successful 3D reconstruction of
the diffraction information through Xnovo Technology
GrainMapper3D™ software yields a completely voxelated
crystallographic representation of the entire sample. The
information recovered from a successful reconstruction
enables extraction of details of the sample’s microstructure, C

such as grains and their morphology, grain boundaries,

grain sizes, grain boundary normals, statistics across the
entire sample, pole figures for texture analysis, etc.
A 3D grain map of a Ti-alloy sample is shown in Figure 3,
where the grains are colored according to their crystallo-
500 µm
graphic orientation in relation to the vertical sample axis
(inverse pole figure). A virtual slice through the sample
is shown in Figure 3b, whereas a measure for the quality Figure 3 A) Visualization of a 3D grainmap of a titanium alloy (Timet 21S) from
reconstruction with the LabDCT GrainMapper3D software. Inverse pole figure
of the LabDCT reconstruction is given to the user through color coding highlights the crystallographic information. B) Virtual cross section
a confidence map as seen in Figure 3c. through the 3D grainmap; scale bar: 100 µm. C) Confidence map of the recon-
struction corresponding to the virtual cross section in B); the gray scale reflects
the confidence value of each voxel from 0% (black) to 100% (white) confidence.
To help validate the accuracy of the LabDCT technique,
comparisons to the closest related technologies, EBSD
and synchrotron DCT and absorption X-ray microscopy
were carried out. In doing so, independent measurements A C

of sample microstructures were obtained. Figure 4 shows

the comparison of a 3D grain structure derived from LabDCT
data (Figure 4a), with the 3D grain structures derived from
high resolution X-ray absorption tomography of an Al-alloy
with segregated Cu on its grain boundaries. Due to the
density differences between Al and Cu the grain boundaries
can be visualized through conventional X-ray absorption B D

tomography. Figure 4b displays the grain structure of the

sample based on segmentation of the absorption contrast
dataset. The 3D grain structures agree remarkably well, with
differences being located at the grain boundaries themselves
as shown through the difference map in Figure 4d. Main
contributing factors to these differences are the segregation 250 µm

of the Cu itself, leading to an uneven distribution and missing

of some grain boundaries (compare Figure 4c); furthermore,
the subsequent segmentation of the grain boundary network
from the absorption tomography reconstruction will introduce
uncertainties in the location of the boundaries.
Figure 4 A) LabDCT reconstruction of Al-Cu sample. B) Volumetric
segmentation from absorption tomography. Colors in A) and B) indicate
a grain index. C) Virtual slice through the absorption tomography in gray-scale.
D) Virtual slice of the difference map between the data of A) and B); the
binarized color scale indicates regions of differences in white and regions
with no difference in black.

39
Characterizing nanomaterials

Technical Note

Also the angular accuracy of the crystallographic orientations

A
were compared to those obtained using synchrotron DCT
and EBSD. Some of the measurements are shown in Figure 5a
along with the corresponding misorientations determined
from the LabDCT measurement. The absolute values of the
misorientation obtained by the two techniques matches
well within approximately one degree. Figure 5b shows a
histogram of the difference in misorientation measured with
EBSD versus LabDCT as well as the difference in misorienta-
tions as measured by LabDCT versus synchrotron DCT.
The DCT techniques – both being X-ray based – differ in
crystallographic orientation by less than 0.3 degrees,
whereas the differences between LabDCT and EBSD
approach one degree consistent with the lower angular
resolution of the EBSD technique [6].

Conclusion B

We have introduced the principles of diffraction contrast

tomography and its application to determining crystalline
grain structure in samples. This illustrates the continued
progress of laboratory XRM to increase the diversity of
imaging modalities that are inspired from synchrotron
Occurrences

origins to solve problems in materials research and related

fields. The unique ZEISS Xradia 620 Versa XRM hardware
architecture enables data acquisition in powerful combina-
tion with advanced reconstruction and analysis capabilities
powered by Xnovo Technology and their experience in
the field of DCT. The continued use and applications
development of this technique will accelerate the way
3D and 4D science is pursued for non-destructively
studying polycrystalline materials. Misorientation Difference [degree]

Figure 5 A) EBSD data from Ti-alloy sample with indications of misorientation

angles as determined by EBSD and LabDCT. B) Histogram of the differences
in mis-orientation as determined by LabDCT vs EBSD, and by LabDCT vs
synchrotron DCT.

References:
[1] S. A. McDonald, et al., Non-destructive mapping of grain orientations in 3D by laboratory X-ray microscopy, Scientific Reports, 5 (2015) 14665.
[2] C. Holzner, et al., Diffraction Contrast Tomography in the Laboratory – Applications and Future Directions, Microscopy Today, July 2016, p. 34-42.
[3] H.F. Poulsen: Three-Dimensional X-ray Diffraction Microscopy (Springer, 2004)
[4] W. Ludwig, P. Reischig, A. King, M. Herbig, E.M. Lauridsen, G. Johnson, T.J. Marrow and J-Y. Buffiere, Rev. Sci. Inst., 80, 033905, (2009)
[5] S.A. McDonald et al., Scientific Reports 7:5151 (2017)
[6] D. Stojakovic. Processing and Applications of Ceramics, 6(1), 1-13, (2012)

40
Characterizing nanomaterials

Technology Note

2 µm

X-ray Nanotomography in the Laboratory

with ZEISS Xradia Ultra 3D X-ray Microscopes
 Characterizing nanomaterials

Technology Note

X-ray Nanotomography in the Laboratory

with ZEISS Xradia Ultra 3D X-ray Microscopes
Author: ZEISS Microscopy

Date: May 2018

X-ray microscopy (XRM) enables non-destructive 3D investigation of a variety of samples across multiple length scales.
Nanoscale XRM with resolution down to tens of nanometers has long been developed at synchrotron radiation facilities,
driven by the development of advanced X-ray optics such as Fresnel zone plates. ZEISS Xradia Ultra is the first instrument
to bring this capability to the user’s own laboratory. In this paper, we discuss the technology behind Xradia Ultra and
resulting performance, including X-ray optics, contrast modes and X-ray energy. The paper concludes with application
areas and upcoming developments.

Xradia Ultra technology

Benefits of laboratory based X-ray nanotomography:
• Non-destructive 3D imaging down to 50 nm resolution
• In situ and 4D imaging: Observe nanostructural
evolution by imaging the same specimen multiple
times under varying conditions, or over time
• Multiscale and correlative imaging: Bridge the gap
between micro-CT and nanometer resolution, but
destructive, electron microscopy techniques
• Contrast for a wide range of materials, from polymers
and tissue to rock, ceramics and metals
On the cover: PEFC Catalyst Layer showing porosity and connected pore space.
Sample courtesy Carnegie Mellon University
Optical architecture
The architecture of Xradia Ultra is conceptually equivalent to an
optical microscope or a transmission electron microscope (TEM).
This architecture is also called transmission X-ray microscope,
or TXM.
• A high-brightness X-ray source is focused onto the specimen
by a high-efficiency capillary condenser
• A Fresnel zone plate objective forms a magnified image of the
specimen on the X-ray camera (detector)
• An optional phase ring can be inserted into the beam path
to achieve Zernike phase contrast to visualize features in
low absorbing specimens
• As the specimen is rotated, images are collected over a range
of projection angles that are then reconstructed into a 3D
tomographic dataset

X-ray Condenser Sample Objective Lens Phase Ring X-ray

Source Lens (Zone plate) Detector

Figure 1: Optical schematic of Xradia Ultra

42
 Characterizing nanomaterials

Technology Note

Capillary condensers
The purpose of the condenser lens in a microscope is to provide
even illumination of the sample across the entire field of view.
Xradia Ultra employs reflective, single bounce capillaries1 for this
purpose that image the X-ray source spot into the sample plane
(see Figure 1).

Reflective capillaries have a number of advantages over Fresnel

zone plates as condenser lenses in laboratory X-ray microscopes,
namely:

• Close to unity focusing efficiency

• Ability to fabricate large numerical aperture (NA) condensers
Figure 2: Capillary condenser
to match NA of high resolution imaging zone plates
• Ability to fabricate large diameter condensers

To prevent excessive blurring of the focal spot, which would

result in a loss of X-ray intensity within the field of view, the
surface roughness inside the capillary must be controlled to
the submicron level.

Fresnel zone plates

Fresnel zone plates are used as microscope objectives to provide
a magnified image of the sample on the X-ray camera. Zone Figure 3: Top: Schematic of a zone plate.
plates are circular diffraction gratings with radially decreasing
Right: SEM image of a zone plate.
line width, acting as a focusing lens for X-rays2,3 (see Figure 3).
Some optical parameters of zone plates are listed in Appendix B.

The spatial resolution that can be achieved, determined by

the Numerical Aperture (NA) in the same way as in an optical
microscope, is about the same as the width of the finest, outer
zones (ΔRn, typically a few tens of nanometers). At the same
time, the zones need to be tall (hundreds of nanometers to more
than a micron) to be efficient. This leads to a requirement for
very high aspect ratio (ratio of height to width) structures.
ZEISS XRM has developed proprietary technology to “stack”
and permanently bond zone plates with nanometer accuracy
to increase the effective zone height and therefore aspect
ratio4 (see Figure 4).

Figure 4: Schematic of the stacking concept,4 where two zone plates are bonded
together to increase the effective thickness of the zones. Zones need to be narrow
(laterally) to achieve high spatial resolution, but thick to achieve high efficiency
(which translates into imaging throughput of the X-ray microscope).

43
 Characterizing nanomaterials

Technology Note

X-ray camera
The X-ray camera in Xradia Ultra is based on a scintillator
optically coupled to a CCD detector. This is the same concept
employed by the Xradia Versa family of submicron resolution
XRMs.

As is illustrated in Figure 5, X-rays are incident on a scintillator,

which converts X-rays to visible light. The visible light image
is then magnified and projected onto a CCD camera by an
optical lens assembly. Applicationimages / Grafics / etc.
Border 0,25px 25% black
Optical performance and spatial resolution
The optical architecture described above achieves spatial
X-ray Scintillator Magnification Camera
resolution below 50 nm. The spatial resolution of an imaging Beam convert to Optics Sensor
visible light
system is the smallest distance two features can be separated
by to still be distinguishable from one another.5Siemens
Siemensstars,
stars,
Figure 5: X-ray camera concept
resolution targets with radially decreasing feature size, are
commonly used in X-ray microscopy to assess resolution
(see Figure 6). Note that spatial resolution is not the same
as pixel size (or voxel size, for 3D imaging). While the voxel
size needs to be smaller than the desired spatial resolution
(it is 16 nm for Xradia Ultra in High Resolution mode at
800X magnification), a small voxel size alone does not
help distinguish features if the optical resolution of the
instrument is insufficient.5

X-ray computed tomography (CT)

One of the unique properties of X-rays is their penetration
power, i.e. their ability to see through and inside optically
opaque, thick specimens. However, a simple two-dimensional
X-ray projection image through a thick specimen with complex
50 nm feature size at tip
internal structure is often not useful because features overlay
each other. Computed tomography (CT), widely used in
the medical field, allows the reconstruction of a 3D virtual
volume from individual projections. More recently, micro- Figure 6: Spatial resolution of Xradia Ultra, demonstrated using a Siemens star
resolution target. The innermost features are 50 nm wide (50 nm lines, 50 nm
and nano-CT6-9
5-8 have pushed this technique to much higher
spaces). Image pixel size is 16 nm.
resolutions.

In CT, many individual projection images of the specimen are

acquired over a range of angles (viewing directions), ideally
covering 180 or 360 degrees. Industrial, micro- and nano-CT
as well as 3D X-ray microscopes typically rotate the specimen
through a stationary X-ray beam for this purpose. This is in
contrast to hospital X-ray CT, where the X-ray source and
detector are rotated around a stationary patient.

44
 Characterizing nanomaterials
Figure 7: X-ray computed tomography illustrated using the example of a shale rock sample. Left: Example projection through the specimen. While some interior features
are visible, the interior structure cannot be determined in detail from a single projection. Center: A projection series over an angular range of 180 degrees was reconstructed
into a virtual volume. The image shows an individual “slice” (virtual cross section) through the specimen. Right: Volume rendering of the reconstructed volume.
The number of projection images acquired is typically in the
hundreds or thousands. Computer algorithms10
9 are
arethen
thenused
used
to reconstruct a virtual, 3-dimensional volume of the specimen.
The most common algorithms are filtered backprojection (FBP)
or Feldkamp-Davis-Krees (FDK, for cone beam CT), although
various advanced (e.g., iterative) algorithms exist and can
yield better results under certain conditions.
The reconstructed volume can then be viewed “slice by slice”
(looking at virtual cross-sections), or visualized in 3D using
various volume rendering methods (see Figure 7). Commonly,
the reconstructed volume is segmented into different compo-
nents based on grayscale or other characteristics. Segmented
volumes can be further analyzed quantitatively in terms of
porosity, tortuosity, particle size distribution or other properties,
or be used as input for advanced modeling algorithms.
The key difference between Xradia Ultra and other micro-
or nano-CT instruments is the use of X-ray optics to achieve
ultrahigh resolution projections as described above.
Absorption and phase contrast
Microscopes may provide a variety of contrast mechanisms
that are sensitive to different properties or components of the
specimen and therefore complement each other. In this section,
we describe absorption and phase contrast, two contrast modes
typically used to visualize morphology of the sample and both
available in Xradia Ultra. Other contrast modes available in other
types of microscopes (light, electron or synchrotron based X-ray
microscopes) include optical or X-ray fluorescence for functional
Technology Note
or elemental imaging, diffraction for crystallographic orientation,
or spectroscopic contrast mechanism for elemental or chemical
imaging.
Absorption contrast
Absorption contrast is essentially shadow contrast, similar to
standard medical X-rays. Material that absorbs strongly shows
up dark in the image, material that absorbs less shows up bright –
like bone vs. tissue in the hospital. Absorption is typically stron-
ger for materials with higher density and/or atomic number.
The mechanism is explained in more detail in the section on
X-ray source energy below.
Absorption is most often used to image pore networks or other
voids (empty space = low/no absorption) in medium to high
density materials, such as rock, calcified tissue, ceramics or
metals. In a multi-phase or composite material, if the compo-
nents are of sufficiently different density, absorption contrast
can also be used to separate the materials (e.g. ceramic-metallic
composites, or cermets).
Phase contrast
10 is primarily used for imaging materials that have
Phase contrast11
low absorption, or to differentiate materials of similar density.
Phase contrast uses the optical effect of refraction rather than
absorption. It is very sensitive to the transition between different
materials (edge enhancement). Note that the term “phase”
here refers to the phase shift of an optical wave, and not to
material phases (like crystallographic phases).
45
 Characterizing nanomaterials

Technology Note

Phase contrast is essential when imaging materials of low

density or atomic number, such as polymers or soft tissue,
since absorption contrast is typically very weak. Figure 8
shows an example of a polymer membrane imaged in
phase contrast.

Due to the edge enhancement properties of phase contrast,

it is also well suited to detect small features close to the resolu-
tion limit, such as fine cracks and defects, in more strongly
absorbing material. It may be used as an initial survey mode to
find small features prior to 3D imaging in absorption contrast. 5 µm
On the other hand, these same edge enhancement properties
mean that different gray scales in the image may not correspond Figure 8: Polymer fibers in a desalination membrane, imaged in phase contrast.
Such low-Z materials are essentially transparent to multi-keV X-rays and therefore
directly to different materials. In such a case, phase contrast may
not able to be visualized in absorption contrast. Sample courtesy of Industrial
be more suitable to detect and visualize small features, whereas Technology Research Institute (ITRI).
absorption contrast is more suitable to quantify and segment
features based on gray scale thresholding. A comparison of
Absorption Contrast Phase Contrast
absorption and phase contrast for a sample of shale rock is
shown in Figure 9.

Multiple implementations of phase contrast imaging have been

developed over time for different microscope architectures.
Xradia Ultra uses the Zernike principle to achieve phase contrast
(see Figure 10). This principle was invented by Frits Zernike in
12 for which he received the
the 1930s for light microscopes,11
13 10 µm 10 µm
Nobel Prize in 1953.12

Figure 9: Comparison of absorption and phase contrast for a shale rock sample.
In absorption contrast (left), the grayscales in the image directly represent different
materials or material phases in the sample, which makes it more suitable for
image segmentation. Phase contrast (right) visualizes fine cracks much more clearly.

Condenser Imaging Phase Ring

Aperture Zone Plate
Undiffracted Light Phase Ring
Image on
Detector

Object

Diffracted Light

Hollow Cone
Illumination

Figure 10: Left: Illustration of Zernike phase contrast. The sample is illuminated by a hollow cone of X-rays. A phase ring is inserted in the back focal plane of the objective
zone plate. The “undiffracted light” (X-rays that have not interacted with the sample) passes through the phase ring and is phase shifted by a quarter wavelength.
“Diffracted light” (X-rays that have interacted with the sample and therefore have been scattered off the direct path) does not pass through the phase ring.
In the detector plane, the diffracted and undiffracted light combine (interfere), which turns phase shifts into intensity variations. Right: SEM image of a phase ring.

46
 Characterizing nanomaterials

Technology Note

X-ray source energy

X-ray Absorption in Material
3D data acquisition times in a laboratory based nanoscale 1.2
XRM are on the time scale of hours. Therefore, optimizing 5.4 keV
1 8 keV
throughput is essential to make the most productive use of the

Relative Intensity
0.8
system. In this section, we will review how the two different
1/e = 37%
X-ray source options, 5.4 keV for Xradia 810 Ultra and 8 keV 0.6

for Xradia 800 Ultra, differ in terms of imaging properties and 0.4

performance for different materials to be imaged. To do so, we

0.2
13 and what that
will review how X-rays get absorbed in matter14
0
means for the resulting image contrast and feature detectability. Propagation Distance (t)
We will explain that in the context of absorption contrast, Absorption length t0

although the concepts apply similarly to phase contrast. For

Figure 11: Schematic illustration of X-ray absorption in matter following the
further details, see Ref. 15, which illustrates contrast, noise Lambert-Beer law. After a propagation distance t0 (which is a function of
and detectability in the context of medical CT. X-ray energy), the intensity drops to 37% of its incident value.

X-ray absorption in matter

X-ray Absorption Length
Absorption of X-rays in matter follows the Lambert-Beer law:
120
Abs. length Al (µm)
100 Abs. length Fe (µm)
I = I0 exp(– t /t 0 )
Absorption Length (µm)

80

where I is the intensity after propagation distance t , I0 is the 60

incident intensity, and t 0 is the absorption length of the material 40

in question at a given X-ray energy. t0 is the distance after which
20
the intensity falls to 1/e = 37% of the incident value. Some-
times, the absorption coefficient μ = 1/t0 is used instead. 0
5000 6000 7000 8000 9000
Photon Energy (eV)

The absorption length is a function of the material, its density

and the X-ray energy. Generally, the absorption length increases
with energy (i.e., harder X-rays are more penetrating), and
decreases with density and atomic number Z (i.e., heaver
materials are more strongly absorbing).
Figure 12: Illustration of X-ray absorption length for two example elements.
Generally, absorption length is longer for lighter elements (aluminum, Al)
compared to heavier elements (iron, Fe). Furthermore, absorption length typically
increases with energy, except for so-called absorption edges, which are
discontinuities in the absorption spectrum. This is illustrated for the example
of iron, which has an absorption edge around 7.1 keV photon energy.

Looking at the two specific X-ray source energies available

for Xradia Ultra, the absorption length is generally longer at Sample 5.4 keV 8.0 keV
8 keV than at 5.4 keV. There are a few exceptions (specifically Iron* 11.7 µm 4.16 µm
vanadium, chromium, manganese, iron and cobalt), which have Aluminum 24.8 µm 77.6 µm
so-called absorption edges (discontinuities in the absorption PMMA 397 µm 1337 µm
length due to the specific atomic energy levels) in between CaCO3 16.5 µm 48.7 µm
the two energies. A few examples are listed in Table 1 below. Silicon 22.4 µm 69.6 µm
To provide for sufficient penetration for imaging, the sample Tungsten 1.14 µm 3.11 µm
thickness should be at most ~2x the absorption length. *Note: Iron is an exception to the rule that 8 keV X-rays can penetrate further
than 5.4 keV X-rays. This is because of the iron X-ray absorption edge at
7.1 keV, above which penetration drops drastically.

Table 1: Approximate X-ray absorption length for select materials. Exact values
will depend on composition and density. Note that X-ray penetration will be larger
if material is porous.

47
 Characterizing nanomaterials

Technology Note

Contrast
Contrast is the difference in light intensity that makes different
features in an image distinguishable from one another. To be
able to detect a feature, it must both be resolved (see Optical
Performance and spatial resolution above) and have sufficiently
strong contrast compared to its surroundings.
As described above, lower energy X-rays are generally absorbed
more strongly and therefore will provide higher contrast. Thus,
as long as transmission remains sufficient, resulting image quality
and/or throughput will be greatly improved at 5.4 keV X-ray
energy compared to 8 keV. However, for materials of higher
density (such as most metals, but note the exception of iron
explained above), or thick specimens, the higher X-ray energy of
8 keV may be needed for sufficient transmission (see Figure 13).
The contrast difference is illustrated on a real world sample in
Figure 14. 5.4 keV X-rays are absorbed more strongly than 8 keV,
resulting in higher contrast and therefore better detectability.

X-ray Penetration: Medium Density Material X-ray Penetration: High Density Material
1.2 1.2
Greater
1 intensity 1
drop equals
Relative Intensity

Relative Intensity

0.8 higher 0.8

contrast
Transmission
0.6 0.6
insufficient
to discern
0.4 0.4 small variations
5.4 keV 5.4 keV
0.2 0.2 8 keV
8 keV

0 0
0 20 40 60 80 100 0 20 40 60 80 100
Penetration Distance (µm) Penetration Distance (µm)

Figure 13: Illustration of 5.4 vs. 8 keV X-ray energy. Left: The greater intensity drop of 5.4 keV X-rays leads to higher contrast (and therefore higher imaging throughput).
Right: If absorption becomes too strong (dense material or sample too thick), transmission at 5.4 keV may be too low to discern small variations, in which case 8 keV is preferred.

Image Contrast

5.4 keV
Image Intensity (a.u.)

8 keV

5.4 keV 8. keV Line Profile Through Image

Figure 14: Contrast difference between 5.4 and 8 keV illustrated using virtual cross-sections of a dentin sample. The lower X-ray energy of 5.4 keV provides stronger
contrast between the bulk intertubular dentin (medium gray), peritubular dentin (light) and tubules (dark). Note that the images follow the convention of displaying
higher density materials in light colors and lower density (air) in dark. The plot on the right shows an intensity profile across the lines indicated in the images.
It is evident that the contrast (intensity difference) is stronger at 5.4 keV than at 8 keV.

48
 Characterizing nanomaterials

Technology Note

Noise
Poisson Distribution for n = 50
Imaging, like any measurement process, introduces noise.
0.06
Noise is a random fluctuation of the measured value around the
0.05
expected, or “true” value. That means, even if we were to image
0.04
a perfectly uniform material, the measured intensity values of

Probability
0.03 —
different pixels (or, the same pixel in multiple measurements) FWHM = 2.35 √50 = 16.6
0.02
would not be exactly the same.
0,01

0
While multiple noise sources exist in an X-ray microscope, we
-0.01
will concentrate here on shot noise, which is intrinsic to the 0 20 40 50 60 80 100

measurement of discrete particles such as X-ray photons. Shot Number of Photons

noise follows a Poisson distribution, which means if we expect

to measure an average number of n photons in many measure-
ments of the same type, the distribution of actual measurements
_
will have a standard deviation (noise) of √n (see Figure 15).
Figure 15: Illustration of Poisson distribution. Assuming we expect to measure a
mean of n=50 photons in many measurements of the same type, the actual number
measured in a single measurement will vary from about 30 to 70 based on the
distribution shown. It can be shown that
_ the full width at half maximum (FWHM)
of this distribution is FWHM = 2.35 √n, or FWHM = 16.6 in this case.

We can then define the Signal-to-Noise Ratio (SNR) as the

ratio of expected number of photons and noise:

_
_
SNR = Signal/Noise = n/√n = √n

In other words, by increasing the exposure time and therefore

counting more X-ray photons, we can decrease the noise level
relative to the measured signal. This is in analogy to photogra-
phy, where in the case of low light, longer exposure times are
used to decrease noise. However, the improvement follows a
square root relationship, i.e. we have to measure four times 3 hours 12 hours
as long to double the SNR.
Figure 16: Virtual cross section of dentin sample from a 3 hr tomogram (left)
and a 12 hour tomogram (right) at 5.4 keV X-ray energy. It is evident that the
This concept is illustrated again for the case of dentin in longer exposure leads to lower noise.

Figure 16. Here, both images were acquired using 5.4 keV
X-ray source energy, meaning the contrast between the
different features is the same. However, it can be seen
that the noise is much lower for the longer exposure.

Putting it all together:

Contrast to Noise Ratio and imaging throughput
So what does all this mean for general image quality and
detectability of features in an image? Ultimately, what counts
is the Contrast-to-Noise Ratio (CNR) of the features of interest
relative to the surroundings. CNR can be increased by either
increasing contrast or decreasing noise. In other words, even
in the case of lower contrast we can detect features by
exposing longer and therefore decreasing noise.

49
 Characterizing nanomaterials

Technology Note

However, if we can increase contrast by using a more suitable • Motion errors of the rotation stage are measured and
X-ray energy, we can achieve sufficient CNR with shorter corrected for by a novel run-out correction system15
16

exposure times, which is generally preferred.
Looking back at Figure 16 again, note that contrast is the same
in both images because they were both acquired at 5.4 keV
X-ray energy. However, CNR is clearly improved with the
12 hour exposure.
Conclusions
We have seen that for most materials, 5.4 keV X-rays are
absorbed more strongly than 8 keV X-rays, leading to higher
contrast and therefore higher imaging throughput. In other
words, equivalent image quality can be achieved with shorter
• The precision motion control system comprises 26 axes
of motion to position and align the X-ray optics, sample,
optical alignment microscope and other components
Application areas
In the following, we list a number of applications where Xradia
Ultra and its predecessor models (Xradia nanoXCT and UltraXRM)
have been used to provide 3D nanostructure data.
Materials research
• Li-ion battery electrodes16-18
17-19

exposure times. If fact, it has been shown that for samples • Polymer electrolyte fuel cell (PEFC) electrodes20-22
19-21

such as rock or calcified tissue, the same contrast to noise • Solid oxide fuel cell (SOFC) electrodes23
22

ratio (image quality) can be achieved about 10 times faster • 3D-ordered macroporous materials for CO2 capture24
23

using a 5.4 keV X-ray source compared to 8 keV. • Osmosis membranes25

24

• Alloys26
25

On the other hand, for samples made of higher-Z (“heavier”) • Superconducting materials27
26

material, 8 keV might be needed to provide sufficient • Nanocomposites28

27

penetration. Ultimately, the preferred source energy varies • Cement28

29

by application. • Subsurface coatings30

29

• High voltage insulators31

30

Furthermore, for a given experiment, it is important to consider

what image quality is sufficient. It may be more productive to Life and environmental science
measure multiple samples at lower CNR in a given time frame • Unstained tissue32
31

than just a single one at higher CNR, as long as the results allow • Nanoparticle uptake in protozoa or soybean roots32,33
33,34

detecting and quantifying the features of interest as required. • Pharmaceutical particles34

35

• Bone and dentin

Mechanical architecture
To achieve 50 nm resolution in a 3D measurement taking several Natural resources
hours, the microscope needs to be mechanically stable over that • Digital rock physics (DRP)36,37
35,36 or general rock

time frame. The following design features are implemented in 38,39 e.g. for carbonate rock, shale rock,
characterization,37,38
Xradia Ultra to achieve that: or tight sand

• Xradia Ultra is built on a low vibration platform to isolate Electronics

the optical system from external vibrations 39,40
• Voids and extrusions in through-silicon via (TSV)40,41
• Careful thermal engineering and calibration prevents or
corrects thermal drifts on the scale of tens of nanometers
between specimen and optics

10

50
 Characterizing nanomaterials

Technology Note

Summary and outlook

Nanoscale 3D X-ray microscopy is the only technique that can
achieve spatial resolution down to 50 nm, non-destructively,
on samples of tens to hundreds of microns in size. As such, it
is particularly well suited to 4D and in situ experiments, where
the same sample is imaged multiple times over time or under
varying conditions, allowing studying microstructural evolution.
Furthermore, nanoscale 3D XRM covers a unique length scale
in a correlative suite of techniques, between micron/submicron
XRM or micro-CT, where larger samples can be studied with
resolution down to one micron or slightly below, and serial
sectioning using electron microscopy, where smaller volumes
can be studied with nanometer resolution, but destructively.
In this White Paper, we described the technology behind
Xradia Ultra, the only laboratory based nanoscale X-ray
microscope. Resolution down to 50 nm is achieved using
X-ray focusing optics such as reflective capillary condensers
and Fresnel zone plate objective lenses. Absorption and phase
contrast modes allow imaging of a wide variety of materials.
Demonstrated applications cover a wide range of examples from
materials research, life and environmental sciences, natural
resources and electronics. Emerging techniques include in situ
imaging, for example under mechanical load, 4D imaging, such
as monitoring corrosion, and correlative imaging across multiple
length scales and using different but complementary techniques.

Appendix A: Why use X-ray optics?

The vast majority of laboratory-based X-ray microscopes and micro CT systems use the principle of cone beam projection through the sample,
followed by X-ray detection on a 2D detector. Rapid developments in the last decade have pushed the achievable spatial resolution down to
about one micron and below. Two concepts are commonly used5(see
(seeFigure
Figure17).
17).

• Conventional micro CT systems typically use a small spot X-ray source and a flat panel detector
Sample with relatively large pixels (~100 µm).
Center of Rotatation
To achieve voxel sizes below one micron, a large geometric magnification (>100x) and therefore very small source-to-sample distance is required.

SourceThe optical magnification stage effectively provides a small

• ZEISS Xradia Versa architecture uses a two-stage (geometric + optical) magnification.
pixel (down to <500 nm) detector. Voxel sizes below one micron can be achieved with low geometric magnification (<10x) and Detector
working
distances of several centimeters.

While both of these concepts keep pushing spatial resolution further below one micron (currently ~600-700 nm), it is ultimately limited by factors
such as achievable X-ray spot size at sufficient power, X-ray detector resolution and practical limitations of acceptable exposure times to achieve
41 that at resolution levels below about 500
sufficient signal-to-noise ratio. It can be shown42 Dss nm, a projection based architecture
Dds Geometricimpractical.
becomes Mag
To achieve spatial resolution higher than that and into the sub-100 nm regime, a lens-based architecture using X-ray focusing optics is required.

Sample Geometric Optical

Center of Rotatation Mag Mag

Source
Detector

Scintillator Detector

Dss Dds Geometric Mag

Figure 17: (Left) Conventional micro CT architecture using a large pixel detector and high geometric magnification to achieve high resolution. This requires a small spot
X-ray source to avoid penumbral blurring, and small source to sample distance (working distance) to achieve small voxel sizes. Resolution degrades quickly as the sample
is moved away from the source. (Right) ZEISS XRM architecture using two-stage magnification. Moderate geometric magnification is followed by switchable optical
Geometric
magnification. This relaxes the Opticaland allows for longer working distance, which is advantageous in the case of interior tomography
need for high geometric magnification
Mag Mag
(imaging a small interior subregion of a large sample at high resolution), and when using in situ sample environments.

11

Scintillator Detector
51
 Characterizing nanomaterials

Technology Note

Appendix B: Fresnel zone plates

The properties of a zone plate can be specified by three parameters such as the diameter D, the outer zone width ΔRn and the wavelength λ.
Then, some other quantities of interest are the number of zones N
N = D/4ΔR
n
the focal length ƒ
ƒ = D ΔRn/λ
and the numerical aperture
NA = λ/2 ΔR
n

Note that the relationship between photon energy E and wavelength λ is given by
E = hc/λ = 1239.842 eV ⋅ nm/λ
where hc = 1239.842 eV ⋅ nm is the product of Planck’s constant h and the speed of light c.

The spatial resolution achievable by a Fresnel zone plate is determined by the outer zone width ΔRn.
Specifically, for uniform plane-wave illumination the transverse resolution δt according to the Rayleigh criterion is given by Abbe’s law:
δt = 0.61λ/NA = 1.22 ΔRn

The depth of focus, or longitudinal resolution δl, is

δl = ± 2ΔRn2/λ

12

52
 Characterizing nanomaterials

Technology Note

Technology Note
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Electrodes Hydrogen Energy 37 (2012),
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8505-8511. DOI: 10.1016/j.ijhydene.2012.02.099
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[22] JR Izzo, et al.,
Epting and“Nondestructive Reconstruction
S. Litster, “Effects and Analysis
of an agglomerate of SOFC Anodes
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Energy 37 (2012),
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JR He,
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al.,“Three-Dimensionally Ordered Macroporous
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H He, et al., “Pore structure
et al., “Three-Dimensionally characterization
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Polymeric Materials Non-destructive
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[25] BM Patterson, et al., “Pore
SS Manickam, “Laboratory micro-
structure and nanoscale
characterization of X-ray tomographic
asymmetric investigation
membranes: of Al–7 at.%Cu
Non-destructive solidification
characterization structures.”
of porosity and tortuosity.”
Mater. Charact.
J. Membr. 95(2014),
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18-26. DOI:
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10.1016/j.memsci.2013.11.044
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BMKim et al., “Microscopic
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Technology Note

[27] V Stepanov et al., “Structural Characterization of RDX-Based Explosive Nanocomposites.” Propell. Explos. Pyrot. 38 (2013), 386-393.
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cement paste.” Cement Concrete Res. 67 (2015), 138-147. DOI: 10.1016/j.cemconres.2014.08.007
[29] NL Cordes et al., “Non-destructive elemental quantification of polymer-embedded thin films using laboratory based X-ray techniques.”
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Spectrochim. Acta B 101 (2014), 320-329. DOI: 10.1016/j.sab.2014.09.016
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[30] R Schurch et al., “Imaging and Analysis Techniques for Electrical Trees using X-ray Computed Tomography.” IEEE T. Dielect. El. In. 21 (2014), 53-63.
DOI: 10.1109/TDEI.2014.6740725
[31] LA Walton, et al., “Morphological Characterisation of Unstained and Intact Tissue Micro-architecture by X-ray Computed Micro- and
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[32] RE Mielke, et al., “Differential Growth of and Nanoscale TiO2 Accumulation in Tetrahymena thermophila by Direct Feeding versus Trophic
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Transfer from Pseudomonas aeruginosa.” Appl. Environ. Microbiol. 79 (2013), 5616-5624. DOI: 10.1128/AEM.01680-13
[33] JH Priester, et al., “Soybean susceptibility to manufactured nanomaterials with evidence for food quality and soil fertility interruption.”
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Proc. Natl. Acad. Sci. U.S.A. 109 (2012), E2451-E2456. DOI: 10.1073/pnas.1205431109
[34] J Wong, et al., “NanoXCT: A Novel Technique to Probe the Internal Architecture of Pharmaceutical Particles.” Pharm. Res. 31 (2014), 3085-3094.
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[35] J Dvorkin et al., “From micro to reservoir scale: Permeability from digital experiments.” The Leading Edge 28 (2009), 1446-1452.
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[42]

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 Characterizing nanomaterials

Technology Note

Evolution of Gemini Electron Optics:

The Next Chapter in Sub-nanometer Imaging Below 1 kV

60 nm
 Characterizing nanomaterials

Authors: Dr. Luyang Han, Dr. Benjamin Tordoff

Carl Zeiss Microscopy GmbH, Oberkochen, Germany
Date: October 2020

Introduction a compound magnetic-electrostatic lens, the Gemini electron

In the last 20 years, a major thrust of research in scanning optics were constantly improved over the decades. In 2015,
electron microscopy (SEM) has been driving the performance ZEISS GeminiSEM 500 was launched, equipped with the Nano-
of systems towards higher resolution at lower voltages. This is twin lens. Its Cs and Cc values were reduced to less than one
motivated by the requirement to further understand surface third of the original Gemini lens design. This design also reduced
structures and their chemistry thanks to breakthroughs in such the magnetic field strength on the sample, which was already
fields as 2D materials, metamaterials and semiconductor device a factor of 1000 less than the sample experiences with an
manufacturing approaching the limit of Moore’s law. immersion lens.

Key to the performance of any SEM, conventional or field 25

emission SEM (FE-SEM), is its electron optical column.

Aberration Coefficients [mm]

ZEISS FE-SEMs are based upon the Gemini column design 20
Cs
which has seen a number of technological advancements
15
since it was first launched in 1993. Low acceleration voltage
imaging is attractive in that it reduces the interaction volume 10
in the specimen, which allows surface sensitive information to
5 Cc
be extracted. Additionally, the charging of insulating samples Cs

by SEM investigation can be avoided by reducing the electron Cc

0
energy used to probe the sample. The reduced radiation 0 5 10 15 20 25 30
damage from low energy SEM imaging is also essential for Beam Energy [KeV]
imaging sensitive specimens such as biological samples and
Figure 1 Spherical and chromatic aberrations (Cs and Cs) of the original Gemini
semiconductor devices [1]. lens (black curve) and the Nano-twin lens (blue curve) with respect to different
beam energies at 1 mm working distance.

Advancements in Objective Lens Designs

It is well known that in order to achieve a fine electron probe New Electron Optical Engine
size at low energy, a special lens design with low spherical ZEISS now introduces several technical achievements to further
and chromatic aberration (Cs and Cc) is necessary. In basic extend the usability and performance of the Gemini column
implementations to overcome this problem, the specimen is design by an improved Nano-twin lens. Moreover, these
immersed in either a strong magnetic field (magnetic single developments achieve better than 1 nm resolution below
pole lens) or a strong electric field (sample bias), or even both. 1 kV without immersing the specimen in an electric or magnetic
However, these methods put severe restrictions on the sample field. Thanks to a new electron optical (EO) engine, the beam
types that can be investigated in addition to their shape and convergence angle is optimized on-the-fly for the current
the ability to tilt such samples. By combining an enclosed working condition to achieve the best possible resolution.
magnetic lens with in-column beam deceleration, the so called The new engine enables a new scanning mode to achieve a very
Gemini design, it is possible to achieve comparable if not better large field-of-view and provides a seamless transition between
resolution than immersion-type lenses, while at the same time sample navigation over large distances and high-resolution
maintaining a high degree of sample flexibility [2] and avoiding imaging. Lastly, the engine delivers extremely rapid automated
immersing the sample in an electromagnetic field. focus and beam alignment using a parallax method.

In 1993 ZEISS developed the first commercial FE-SEM, called Examples of high-resolution images obtained at low beam
the DSM-982 with Gemini electron optics based on such a energy are shown in the following figures. The montmorillonite
principle. A distinct characteristic of this compound lens is that particles shown in Fig. 2 are non-conducting. To avoid
both chromatic and spherical aberrations (Cc and Cs) decrease charging, this specimen is imaged at 800 V using 15 pA current
with electron energy (Fig. 1), which is especially helpful to by selecting a corresponding aperture size. The new engine
achieve good resolution at very low beam energies, such as automatically optimizes the condenser excitation to achieve the
those lower than 1 keV. The DSM-982 was the first commercial best resolution at this specific condition. Since such a sample
SEM to demonstrate resolution better than 10 nm with less is not conductive, it is not feasible to apply a potential on the
than 1 keV electron energy [3]. Based on the same principle of specimen, as this would cause electrostatic charging effects

2
56
 Characterizing nanomaterials
which could damage the system. Thankfully such a potential is
not required to achieve an image of exceptional resolution with
the Gemini column design combined with Nano-twin lens and
the new engine. Similarly, the FeMn magnetic nanoparticles
shown in Fig. 3 cannot be immersed in a magnetic field for to help navigate across the specimen. This is achieved by a
imaging as this would attract these nanoparticles towards the
lens, possibly causing damage to the SEM system. Again, the
Gemini column design with Nano-twin lens and new EO engine
circumvent this problem through the well know Gemini optical
design principle.
200 nm
Figure 2 High resolution image of montmorillonite particles obtained at
800 V beam energy. Even though the sample is non-conductive, fine flakes on
the fractured surface can be well resolved.
60 nm
Figure 3 High resolution image of FeMn magnetic nanoparticles obtained at
1 kV beam energy. Even though the sample is magnetic, the highest resolution
can still be achieved.
While resolution is usually treated as the benchmark of any SEM,
the importance of lower magnification performance is usually
underappreciated. The new EO engine from ZEISS provides
a novel scanning mode to achieve a large field of view (FoV)
different arrangement of the scanning system, while maintaining
the beam energy and lens configuration unchanged.
This assures fast transition between the normal imaging mode
and large field of view mode without changing the voltage of
any electron-optic components, and all detectors in the FE-SEM
system maintain a consistent contrast and performance. In the
example shown in Fig. 4, three Euro coins are imaged at 5 kV
beam energy. In normal imaging mode the maximum FoV is
about 4 mm, only enough to cover a small portion of a single
coin. The new EO engine drives the system to allow all three
coins to be imaged in the same frame.
1 mm
5 mm
Figure 4 Euro coins imaged at maximum FoV in normal imaging mode (upper)
and in high FoV mode (lower) using the new EO engine. The blue square shows
the position and size of maximum FoV in normal imaging mode.
3
57
 Characterizing nanomaterials

Another advantage of the new EO engine can be seen in the Conclusion

resulting auto functions for fine focusing and beam alignment. After pioneering the development of low kV imaging without
Here EO calculations are used to adjust focus and align the using immersion lenses, ZEISS has continued to innovate in
microscope. The time to result is shown in Table 1 showing a this field with the introduction of the Nano-twin lens in 2015.
particularly fast autofocusing time. As an example, in Fig. 5 the The latest development in this field is the new EO optical
precision of the fine autofocus is shown in an image pair, before engine. This EO engine provides further increases in resolution
and after autofocusing. This new capability compliments the at low kV by optimizing the convergence angle of electrons
coarse-autofocus, auto-stigmation and auto-contrast brightness travelling through the column at given conditions. The engine
functions available today. With the design of various degrees also substantially increases the available field of view using the
of automation, the improved speed and ease of use will be high-performance Nano-twin lens. System ease of use is further
available to all users independent of their skills. enhanced through the engine’s new autofocus and auto beam
alignment functions. Notably, the new autofocus capability of
Duration the system brings the operation into the millisecond domain for
Auto focus <1s the first time.
Auto align <4s

Table 1 Performance of EO engine based auto functions.

10 µm

after 1 sec focusing

10 µm

Figure 5 Example of ultra-fast fine focus.

Reference:
[1] Imaging formation in Low-voltage scanning electron microcopy, L. Reimer, SPIE optical engineering press, 1993
[2] Handbook of Charged Particle Optics (2nd edition), J. Orloff, CRC Press, 2009
[3] H. Jaksch, J. P. Martin, Fresenius, Journal of Analytical Chemistry October 1995, Volume 353, Issue 3-4, pp 378-382
High-resolution, low-voltage SEM for true surface imaging and analysis

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 Characterizing nanomaterials

Application Note

Investigating Sweet Spot Imaging of Perovskite

Catalysts Bearing Exsolved Active Nanoparticles
 Characterizing nanomaterials

Authors: Maadhav Kothari and Andy Holwell

Carl Zeiss Microscopy Ltd, Cambridge, UK
Professor John Irvine
University of St Andrews, St Andrews, UK
Date: February 2022

Abstract
Platinum (Pt), generally dispersed on a solid oxide support,
has been widely used for catalytic chemical reactions in
automobile, chemical refining, and energy industries.
During the reactions, Pt is exposed to severe conditions,
e.g., high temperature events and impurities, that cause
Pt agglomeration and poisoning, respectively, resulting
in activity/stability losses. Perovskite materials are designed
with Pt for significant catalytic properties through novel
doping and exsolution methods 1.
1 µm

In order to accurately determine the catalytic ability of

Pt nanoparticles, it is important to understand the structure
and morphology of nanoparticles. Typical scanning electron
microscopy methods do not reveal the morphological
characteristics of nanoparticles due to the lack of electron
beam stability. Here we demonstrate imaging techniques
employed to accurately determine particle size and morphology.

This method can improve the catalytic analysis of Pt loading,

size, dispersion, and active sites determination.
400 nm
Main
Recent advances in new energy and heterogenous catalytic
materials have resulted in novel ceramics being produced for
a wide array of applications such as fuel cells, autocatalysis,
and chemical feedstock production 2,3,4. One such material
is non-stoichiometric A-site deficient perovskite, with a
100 nm
catalytically active metal doped on the B site. It is possible
through controlled synthesis and reduction conditions to Figure 1 Distribution of platinum decoration on reduced areas of perovskite
tailor the size and morphology of nanoparticles through
emergence of the active metal cations.

To study the distribution, size, shape and morphology,

careful scanning electron microscopy (SEM) needs to be
deployed. The perovskite ceramic sensitivity, non-conductive
nature, and nanoparticle distribution on the reduced area
results in a difficult to image surface. This can often result in
nano decoration being missed altogether due to lack of material
understanding and microscopic technique.

2
60
 Characterizing nanomaterials
Often in industrial testing for autocatalysis, a pelletized
and sieved sample is observed and tested. This can lead to
significant imaging issues due in part to debris, carbonaceous
deposits from catalytic testing, and fine static particulate
matter that can significantly reduce the sharpness of a
micrograph. Typically, a solution to non-conductive materials
would be sputter coat with either Gold (Au), Platinum (Pt) or
Carbon (C) or a combination of coats ranging from 20 nm
to 5 µm thickness for ceramic type materials. This can be
counterproductive as often coating can lead to a masking of
nanoparticles and false identification of both nanoparticles and
carbonaceous deposits. Another factor in correctly identifying
nanoparticles and distribution is the ability of substrate ceramic
to be coated and catalytically react with typical desorption/
adsorption-based techniques to characterize active metal sites,
rendering these methods ineffective. SEM image analysis of
nanoparticles is therefore a major area of interest to determine
activity, stability, dispersion, and morphological features.
Electron beam sensitive materials often require additional
expertise and knowledge of the SEM best practice imaging
technique known as sweet spot 5 imaging. Here we demon-
strate analytical micrographs of nanoparticles emerging
from reduced La 0.4 Ca 0.3925Ba 0.0075Pt 0.005Ti 0.995O3-δ 1 taken as the next step. Finally, an approximate aperture is required
noted as Pt+LCT.
Results and Discussion
A-site deficient perovskite Pt+LCT was synthesized using solid
state synthesis and then reduced under 5% H2 /Ar resulting in 0.5
wt% Pt. Emerged nanoparticles on the surface were studied using
ZEISS Sigma 300, ZEISS Sigma 500, ZEISS GeminiSEM 360 and
ZEISS GeminiSEM 560. For determining imaging conditions
of Pt+LCT a range of apertures, working distance from beam
Microscope keV imaged Aperture µm SE / Inlens
Sigma 300 10,5,3,1 50, 20, 15,10 SE & Inlens SE
Sigma 500 10,5,3,1 50, 20, 15,10 SE & Inlens SE
GeminiSEM 360 10,5,3,1 50, 20, 15,10 SE & Inlens SE
GeminiSEM 560 10,5,3,1-<1 50, 20, 15,10 SE & Inlens SE
Table 1 Range of conditions used to determine the sweet spot of the perovskite catalysts
pole and detector choice were selected to facilitate the
best imaging practice (table 1). A range of detectors available
on ZEISS field emission scanning electron microsopes (FE-SEM)
was chosen, all using the ZEISS Gemini electron optical column.
For ceramic type materials and non-conductive materials
it is best practice to image either at low keV (<5 keV) or
variable pressure with a low working distance in relation
to the pole piece. This is often different for individual
microscopes, especially if the detectors and beam stability
are variable. To find the best practice imaging for a certain
material, a sweet spot analysis is required where various
parameters are run through stepwise to find the best possible
imaging conditions. By running through the test matrix in
Table 1 the ideal conditions for the ZEISS GeminiSEM family
columns were found.
It is clear that imaging ceramics at a higher keV (above 5 keV)
damages the surface and sub-surface, resulting in degradation
of the material within the chamber. An area of non-interest is
initially advised to be selected for use with higher keV (5-20 keV).
Once an optimal keV is chosen for the solution, tailoring the
working distance for advanced resolution is advised to be under-
between 5 µm of the selected aperture, e.g., 10 µm-25 µm
should be experimented with to find the optimal conditions and
increase the brilliance of imaging. Imaging snapshots can be
taken with a rapid line scan initially to acquire the appropriate
imaging settings in combination with drift correction. Further
image corrections can be made post scanning with the
softwares ZEISS SmartSEM and ZEISS SmartSEM Touch. Once
imaging conditions have been selected, it is possible to observe
regions of interest without damaging the selected area.
VP/BSE Ideal condition Example
VP-BSE 30 Pa 5 keV, 20 µm aperture, Inlens SE Figure 3
Working distance: 3.5 mm
VP-BSE, Inlens 3 keV, 20 µm aperture, Inlens SE Figure 4
Working distance: 3-5 mm
VP-BSE 20-30 Pa 1-3 keV 10-20 µm aperture, Figure 5
Inlens SE and VP-BSE 20 Pa
Working distance: 4-7 mm
Inlens EsB 1-3 keV 10-20 µm aperture, Figure 6
Inlens SE and Inlens EsB
Working distance: 2-7 mm
3
61
 Characterizing nanomaterials

1 µm 500 nm

Figure 2 Working distance (2.5 mm) needed for imaging nanoparticles on ceramic Figure 3 (L) Standard SE detector with no decoration visible;
(R) use of Inlens SE technology allows nanoparticle decoration to be observed

It was found that for perovskite an aperture of 20 µm was

sufficient for imaging and a working distance of less than 3 mm
(Figure 2) across the Sigma family was required to see nano
meter scaled features on the surface of the materials.

Smaller nanoparticles of <8 nm finely dispersed on the

surface of the perovskite were not completely visible with the
secondary electron detector (ETSE); this can often be misleading
for microscopists who can miss nanoparticle decoration by
using the wrong detector. By using the Inlens SE detector, full
600 nm
nanoparticle decoration was visible. Images were taken using
a rapid <4s line scan as any longer would result in visible
beam damage to the material, as can be seen in Figure 3. 1 2

Further image correction and processing can be tailored

automatically for the Sigma family. SmartSEM Touch
200 nm
can be used alongside imaging (Figure 4) to significantly
reduce time spent correcting image sharpness as well as
brightness and contrast. 3 4

For GeminiSEM 360 and GeminiSEM 560 microscopes,

a magnetic-clamp Kline stage locking the eucentric stage
in place was used to further stabilize the imaging as well
as the pendulum-based motion stabilization for the core
Figure 4 SmartSEM touch allows for rapid image improvements with automated
of the microscope. This enables a lower working distance drift correction and auto brightness contrast sharpening
from the pole piece and enables imaging of unstable or
non-anchored particles to the stub (Figure 5).

50 µm 400 nm

Figure 5 Non-anchored particles can be easily observed due to the stability provided by the GeminiSEM family magnetic clamp,
rendering stage movement minimal and improving image quality

4
62
 Characterizing nanomaterials

2 µm 200 nm

2 µm 1 µm

Figure 6 Inlens BSD allows for greater contrast and sub surface morphology Figure 7 The use of variable pressure mode allows for delicate imaging in both
identification of LCCNT Inlens SE, SE and BSE modes with the Sigma series

Further analytical imaging can be achieved with energy Conclusions

selective backscattered detector (Inlens EsB). Subsurface
nanoscale composition is made visible with a clear compo-
sitional contrast. The second annular incolumn detector is
located above the Inlens SE detector allows for more detailed
material information and contrast in Figure 6.
Pt nanoparticles are visible in NanoVP variable pressure
mode with a back scatter detector (BSD), Figure 7, and can
provide structurally significant information through contrast
and subsurface morphology. Further enhancement can be
achieved at variable pressure alongside a low working
distance relative to the pole piece (between 3-5 mm);
nanoparticles are visible using VP mode. Significantly,
Inlens detectors can be used in VP mode and therefore enhance
imaging and provide contrast alongside BSD images.
Sweet spot imaging conditions can be determined for the
ZEISS FE-SEM range (in this case Sigma and GeminiSEM). It plays
a practical and methodical role in finding optimal conditions
for imaging of samples. For ceramics such as the ones used in
this study, catalytically active nanoparticles decorating an A-site
deficient perovskite, careful imaging and analysis are major
tools in determining catalytic activity. It was found that sweet
spot imaging conditions were not dissimilar between the
ZEISS Sigma and GeminiSEM families with subtle differences
for each microscope type. The stability and stage of the
ZEISS GeminiSEM family were found to increase image
stabilization and brilliance, resulting in high resolution imaging
at low keV. Alongside this, Inlens SE technology was found to
be the most important in observing nanoparticle decoration
for both Sigma and GeminiSEM. Further detectors can be used
in parallel such as Inlens EsB to provide more subsurface and
morphologically distinct regions of interest.

References
[1] Kothari, M. et al. Platinum incorporation into titanate perovskites to deliver emergent active and stable platinum nanoparticles.
Nat. Chem. 13, 677–682 (2021).
[2] Cassidy, M., Gamble, S. & Irvine, J. Application of Exsolved Structures as a Route to More Robust Anodes for Improved Biogas Utilisation
in SOFCs. ECS Trans. (2015).
[3] Papargyriou, D., Irvine, J.T.S. Nanocatalyst exsolution from (La, Sr)(Cr, M, Ni) O 3 (M Mn, Fe) perovskites for the fuel oxidation layer
of Oxygen Transport Membranes. Solid State Ionics (2015).
[4] Neagu, D., Oh, T., Miller, D., Ménard, H. & Bukhari, S. Nano-socketed nickel particles with enhanced coking resistance grown in situ by
redox exsolution. Nat. Commun. (2015).
[5] Tandokoro, K., Nagoshi, M., Kawano, T., Sato, K. & Tsuno, K. Low-voltage SEM contrasts of steel surface studied by observations and
electron trajectory simulations for GEMINI lens system. Microscopy 67, 274–279 (2018).

5
63
 07745 Jena, Germany
microscopy@zeiss.com
www.zeiss.com/microscopy
Carl Zeiss Microscopy GmbH
Characterizing nanomaterials

64
Not for therapeutic use, treatment or medical diagnostic evidence. Not all products are available in every country. Contact your local ZEISS representative for more information.
CZ 06-2023 | Design, scope of delivery and technical progress subject to change without notice. | © Carl Zeiss Microscopy GmbH
Not for therapeutic use, treatment or medical diagnostic evidence. Not all products are available in every country. Contact your local ZEISS representative for more information.
EN_42_013_352 | CZ 02-2022 | Design, scope of delivery and technical progress subject to change without notice. | © Carl Zeiss Microscopy GmbH
Characterizing nanomaterials

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