ISO 19749:2021
(Main)Nanotechnologies — Measurements of particle size and shape distributions by scanning electron microscopy
Nanotechnologies — Measurements of particle size and shape distributions by scanning electron microscopy
This document specifies methods of determining nanoparticle size and shape distributions by acquiring and evaluating scanning electron microscope images and by obtaining and reporting accurate results. NOTE 1 This document applies to particles with a lower size limit that depends on the required uncertainty and on the suitable performance of the SEM, which is to be proven first -according to the requirements described in this document. NOTE 2 This document applies also to SEM-based size and shape measurements of larger than nanoscale particles.
Nanotechnologies — Détermination de la distribution de taille et de forme des particules par microscopie électronique à balayage
Le présent document spécifie des méthodes permettant de déterminer les distributions de taille et de forme des nanoparticules, par l’acquisition et l’évaluation d’images obtenues avec un microscope électronique à balayage, puis l’obtention de résultats exacts et la rédaction de rapports. NOTE 1 Le présent document s’applique aux particules dont la limite de taille inférieure dépend de l’incertitude exigée et des performances appropriées du MEB, après démonstration de sa conformité aux exigences décrites dans le présent document. NOTE 2 Le présent document s’applique également aux mesurages par MEB de taille et de forme des particules de taille supérieure à l’échelle nanométrique.
General Information
Buy Standard
Standards Content (Sample)
INTERNATIONAL ISO
STANDARD 19749
First edition
2021-07
Nanotechnologies — Measurements of
particle size and shape distributions
by scanning electron microscopy
Nanotechnologies — Détermination de la distribution de taille et de
forme des particules par microscopie électronique à balayage
Reference number
ISO 19749:2021(E)
©
ISO 2021
---------------------- Page: 1 ----------------------
ISO 19749:2021(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2021
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting
on the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address
below or ISO’s member body in the country of the requester.
ISO copyright office
CP 401 • Ch. de Blandonnet 8
CH-1214 Vernier, Geneva
Phone: +41 22 749 01 11
Email: copyright@iso.org
Website: www.iso.org
Published in Switzerland
ii © ISO 2021 – All rights reserved
---------------------- Page: 2 ----------------------
ISO 19749:2021(E)
Contents Page
Foreword .v
Introduction .vi
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 2
3.1 General terms . 2
3.2 Core terms: image analysis . 4
3.3 Core terms: statistical symbols and definitions . 4
3.4 Core terms: measurands and descriptors . 6
3.5 Core terms: metrology . 8
3.6 Core terms: scanning electron microscopy .10
4 General principles .11
4.1 SEM imaging .11
4.2 SEM image-based particle size measurements .12
4.3 SEM image-based particle shape measurements .13
5 Sample preparation .13
5.1 Sample preparation fundamental information .13
5.2 General recommendations.14
5.3 Ensuring good sampling of powder or dispersion-in-liquid raw materials .14
5.3.1 Powders .14
5.3.2 Nanoparticle dispersions in liquids .15
5.4 Ensuring representative dispersion .15
5.5 Nanoparticle deposition on a substrate .15
5.5.1 General.15
5.5.2 Nanoparticle deposition on wafers and chips of silicon or other materials .16
5.5.3 Nanoparticle deposition on TEM grids .17
5.6 Number of samples to be prepared .18
5.7 Number of particles to be measured for particle size determination .18
5.8 Number of particles to be measured for particle shape determination .19
6 Qualification of the SEM for nanoparticle measurements .19
7 Image acquisition .19
7.1 General .19
7.2 Setting suitable image magnification and pixel resolution .23
8 Particle analysis .24
8.1 Particle analysis fundamental information .24
8.2 Individual particle analysis .25
8.3 Automated particle analysis .25
8.4 Automated particle analysis procedure example .26
9 Data analysis .27
9.1 General .27
9.2 Raw data screening: detecting touching particles, artefacts and contaminants .27
9.3 Fitting models to data .27
9.4 Assessment of measurement uncertainty .27
9.4.1 General.27
9.4.2 Example: Measurement uncertainty for particle size measurements .28
9.4.3 Bivariate analysis .29
10 Reporting the results .29
Annex A (normative) Qualification of the SEM for nanoparticle measurements .31
Annex B (informative) Cross-sectional titanium dioxide samples preparation .36
© ISO 2021 – All rights reserved iii
---------------------- Page: 3 ----------------------
ISO 19749:2021(E)
Annex C (informative) Case study on well-dispersed 60 nm size silicon dioxide nanoparticles .38
Annex D (informative) Case study on 40 nm size titanium dioxide nanoparticles .46
Annex E (informative) Example for extracting particle size results of SEM-based
nanoparticle measurements using ImageJ .55
Annex F (informative) Effects of some image acquisition parameters and thresholding
methods on SEM particle size measurements .57
Annex G (informative) Example for reporting results of SEM-based nanoparticle measurements 61
Bibliography .71
iv © ISO 2021 – All rights reserved
---------------------- Page: 4 ----------------------
ISO 19749:2021(E)
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out
through ISO technical committees. Each member body interested in a subject for which a technical
committee has been established has the right to be represented on that committee. International
organizations, governmental and non-governmental, in liaison with ISO, also take part in the work.
ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of
electrotechnical standardization.
The procedures used to develop this document and those intended for its further maintenance are
described in the ISO/IEC Directives, Part 1. In particular, the different approval criteria needed for the
different types of ISO documents should be noted. This document was drafted in accordance with the
editorial rules of the ISO/IEC Directives, Part 2 (see www .iso .org/ directives).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
any patent rights identified during the development of the document will be in the Introduction and/or
on the ISO list of patent declarations received (see www .iso .org/ patents).
Any trade name used in this document is information given for the convenience of users and does not
constitute an endorsement.
For an explanation of the voluntary nature of standards, the meaning of ISO specific terms and
expressions related to conformity assessment, as well as information about ISO's adherence to the
World Trade Organization (WTO) principles in the Technical Barriers to Trade (TBT), see www .iso .org/
iso/ foreword .html.
This document was prepared by Technical Committee ISO/ TC 229, Nanotechnologies.
Any feedback or questions on this document should be directed to the user’s national standards body. A
complete listing of these bodies can be found at www .iso .org/ members .html.
© ISO 2021 – All rights reserved v
---------------------- Page: 5 ----------------------
ISO 19749:2021(E)
Introduction
This document provides guidance for measuring and reporting the size and shape distributions of
nanometer-scale particles using images acquired by the scanning electron microscope (SEM). This
document applies to the SEM-based measurement of larger particles also. Nanoparticles are three-
dimensional (3D) objects, but the SEM image is only a two-dimensional (2D) representation of the 3D
shape from a certain viewing angle. The SEM image carries valuable information about the size and
shape of particles. While the SEM image does contain a certain amount of 3D information, for sake of
simplicity, this document does not deal with reconstructing 3D information. Rigorous three-dimensional
characterization of nanoparticles would include size, shape, surface structure (e.g. texture), surface
and internal material composition, and their locations in the investigated 3D volume. This document
deals with two attributes of morphology, size and shape, for discrete and aggregated nano-objects
(materials with at least one dimension in the nanometer-scale, i.e. within 1 nm to 100 nm). Suitable
sample preparation is essential to obtaining high-quality electron microscope images and preferred
techniques often vary with the sample material. It is equally important to make sure that the SEM itself
is suitable to carry out the measurements with the required uncertainty. Typical guidance suggests that
a large number, several hundreds or thousands of particles need to be measured for statistically sound
size and shape distribution results. The actual number of nano-objects needed to be measured depends
on the sample, the required uncertainty and on the performance of the SEM. Statistical evaluation of
the data and the evaluation of uncertainty of the measurands are included as part of the measurement
and reporting procedures.
This document contains measurement procedures, particle and data analysis and reporting clauses. In
the Annexes, there are specific examples for measurements and guidance for the qualification of the
SEM for reliable quantitative measurements. Automation of the image acquisition and data analysis can
reduce cost and improve the quality of the results. Measurements of samples of discrete nanoparticles
are generally easier to carry out with automated image acquisition and particle analysis systems.
Measurements of complex discrete nanoparticles, and aggregates or agglomerates of nanoparticles
may require operator-assisted image acquisition and analysis. Evaluation of particle shape is facilitated
by many pertinent analysis software solutions that allow for automatic selection of various shape
attributes as well.
vi © ISO 2021 – All rights reserved
---------------------- Page: 6 ----------------------
INTERNATIONAL STANDARD ISO 19749:2021(E)
Nanotechnologies — Measurements of particle size and
shape distributions by scanning electron microscopy
1 Scope
This document specifies methods of determining nanoparticle size and shape distributions by acquiring
and evaluating scanning electron microscope images and by obtaining and reporting accurate results.
NOTE 1 This document applies to particles with a lower size limit that depends on the required uncertainty
and on the suitable performance of the SEM, which is to be proven first -according to the requirements described
in this document.
NOTE 2 This document applies also to SEM-based size and shape measurements of larger than nanoscale
particles.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
ISO/IEC Guide 99, International vocabulary of metrology — Basic and general concepts and associated
terms (VIM)
ISO 9276-1, Representation of results of particle size analysis — Part 1: Graphical representation
ISO 9276-2, Representation of results of particle size analysis — Part 2: Calculation of average particle
sizes/diameters and moments from particle size distributions
ISO 9276-3, Representation of results of particle size analysis — Part 3: Adjustment of an experimental
curve to a reference model
ISO 9276-5, Representation of results of particle size analysis — Part 5: Methods of calculation relating to
particle size analyses using logarithmic normal probability distribution
ISO 9276-6, Representation of results of particle size analysis — Part 6: Descriptive and quantitative
representation of particle shape and morphology
ISO 13322-1, Particle size analysis — Image analysis methods — Part 1: Static image analysis methods
ISO 16700, Microbeam analysis — Scanning electron microscopy — Guidelines for calibrating image
magnification
ISO/IEC 17025, General requirements for the competence of testing and calibration laboratories
ISO/TS 24597:2011, Microbeam analysis — Scanning electron microscopy — Methods of evaluating image
sharpness
ISO 26824, Particle characterization of particulate systems — Vocabulary
ISO/TS 80004-1, Nanotechnologies — Vocabulary — Part 1: Core terms
ISO/TS 80004-2, Nanotechnologies — Vocabulary — Part 2: Nano-objects
ISO/TS 80004-3, Nanotechnologies — Vocabulary — Part 3: Carbon nano-objects
ISO/TS 80004-4, Nanotechnologies — Vocabulary — Part 4: Nanostructured materials
© ISO 2021 – All rights reserved 1
---------------------- Page: 7 ----------------------
ISO 19749:2021(E)
ISO/TS 80004-6, Nanotechnologies — Vocabulary — Part 6: Nano-object characterization
3 Terms and definitions
For the purposes of this document, the terms and definitions given in ISO/IEC Guide 99, ISO 9276-6,
ISO 26824, ISO/TS 80004-1, ISO/TS 80004-2, ISO/TS 80004-3, ISO/TS 80004-4, ISO/TS 80004-6, and
the following apply.
ISO and IEC maintain terminological databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https:// www .iso .org/ obp
— IEC Electropedia: available at http:// www .electropedia .org/
3.1 General terms
3.1.1
nanoscale
length range from approximately 1 nm to 100 nm
Note 1 to entry: Properties that are not extrapolations from larger sizes are predominantly exhibited in this
length range.
[SOURCE: ISO/TS 80004-1:2015, 2.1]
3.1.2
nano-object
discrete piece of material with one, two or three external dimensions in the nanoscale (3.1.1)
[SOURCE: ISO/TS 80004-1:2015, 2.5, modified — Note 1 to entry and the source have been deleted.]
3.1.3
particle
minute piece of matter with defined physical boundaries
[SOURCE: ISO/TR 16197:2014, 3.10, modified — Notes 1, 2 and 3 to entry and the source have been
deleted.]
3.1.4
primary particle
original source particle (3.1.3) of agglomerates (3.1.5) or aggregates (3.1.6) or mixtures of the two
[SOURCE: ISO 26824:2013, 1.4, modified — Notes 1, 2 and 3 to entry have been deleted.]
3.1.5
agglomerate
collection of weakly or medium strongly bound particles (3.1.3) where the resulting external surface
area is similar to the sum of the surface areas of the individual components
Note 1 to entry: Agglomerate originates from the Latin “agglomerare” meaning “to form into a ball”.
Note 2 to entry: The forces holding an agglomerate together are weak forces, for example van der Waals forces or
simple physical entanglement.
Note 3 to entry: Agglomerates are also termed secondary particles and the original source particles are termed
primary particles (3.1.4).
[SOURCE: ISO 26824:2013, 1.2, modified — Note 1 to entry has been added.]
2 © ISO 2021 – All rights reserved
---------------------- Page: 8 ----------------------
ISO 19749:2021(E)
3.1.6
aggregate
particle (3.1.3) comprising strongly bonded or fused particles where the resulting external surface area
is significantly smaller than the sum of surface areas of the individual components
Note 1 to entry: The forces holding an aggregate together are strong forces, for example covalent bonds, or those
resulting from sintering or complex physical entanglement, or otherwise combined former primary particles
(3.1.4).
Note 2 to entry: Aggregate comes from the Latin “aggregat” meaning “herded together”.
Note 3 to entry: Figure 1 shows examples of individual, aggregate and agglomerate (3.1.5) particles.
NOTE The images are projected views from certain angles of the 3D objects. Depending on the viewing
angle, the observable size of particles can vary substantially.
Figure 1 — SEM images of individual gold (left) and carbon black aggregate (middle) and
corundum agglomerate (right) particles
[SOURCE: ISO 26824:2013, 1.3, modified — Notes 2 and 3 to entry have been added.]
3.1.7
nanoparticle
nano-object (3.1.2) with all external dimensions in the nanoscale (3.1.1) where the lengths of the longest
and shortest axes of the nano-object do not differ significantly
[SOURCE: ISO/TS 80004-2:2015, 4.4, modified — Note 1 to entry has been deleted.]
3.1.8
particle size
x
dimension of a particle (3.1.3) determined by a specified measurement method and under specified
measurement conditions
Note 1 to entry: Different methods of analysis are based on the measurement of different physical properties.
Independent of the particle property actually measured, the particle size can be reported as a linear dimension,
an area, or a volume.
Note 2 to entry: The symbol x is used denote linear particle (3.1.3) size. However, it is recognized that the symbol
d is also widely used. Therefore, the symbol x may be replaced by d.
3.1.9
particle size distribution
distribution of the quantity of particles (3.1.3) as a function of particle size (3.1.8)
[SOURCE: ISO/TS 80004-6:2021, 4.1.2, modified — Notes 1 and 2 to entry have been deleted.]
© ISO 2021 – All rights reserved 3
---------------------- Page: 9 ----------------------
ISO 19749:2021(E)
3.1.10
particle shape
external geometric form of a particle (3.1.3)
Note 1 to entry: Shape description requires two scalar descriptors, i.e. length and breadth.
[SOURCE: ISO/TS 80004-6:2021, 4.1.3, modified — Note 1 to entry has been added.]
3.1.11
analytical sample
portion of material, resulting from the original sample or composite sample by means of an appropriate
method of sample pretreatment and having the size (volume/mass) necessary for the desired testing or
analysis
Note 1 to entry: The sample in analytical chemistry is a portion of material selected from a larger quantity of
material. The term needs to be qualified, for example, bulk sample, representative sample, primary sample,
bulked sample, test sample. The term 'sample' implies the existence of a sampling error, i.e. the results obtained
on the portions taken are only estimates of the concentration of a constituent or the quantity of a property present
in the parent material. If there is no or negligible sampling error, the portion removed is a test portion, aliquot
or specimen. The term 'specimen' is used to denote a portion taken under conditions such that the sampling
variability cannot be assessed (usually because the population is changing), and is assumed, for convenience, to
be zero. The manner of selection of the sample should be prescribed in a sampling plan.
[SOURCE: ISO 11074:2015, 4.1.3, modified — Note 1 to entry has been added.]
3.2 Core terms: image analysis
3.2.1
binary image
digitized image consisting of an array of pixels (3.2.2), each of which has a value of 0 or 1, whose values
are normally represented by dark and bright regions on the display screen or by the use of two distinct
colors
[SOURCE: ISO 13322-1:2014, 3.1.2]
3.2.2
pixel
smallest element of an image that can be uniquely processed, and is defined by its spatial coordinates
and encoded with colour values
[SOURCE: ISO 12640-2:2004, 3.6, modified — Note 1 to entry has been deleted.]
3.2.3
pixel resolution
number of imaging pixels (3.2.2) per unit distance of the detector
Note 1 to entry: The typical unit is sometimes expressed as dots per inch (dpi).
[SOURCE: ISO 29301:2017, 3.24, modified — the hyphen has been deleted in this term.]
3.3 Core terms: statistical symbols and definitions
3.3.1
arithmetic mean
sum of values divided by the number of values
Note 1 to entry: See ISO 9276-1:1998 for other quantity measures and types of distributions.
4 © ISO 2021 – All rights reserved
---------------------- Page: 10 ----------------------
ISO 19749:2021(E)
3.3.2
standard deviation
measure of the dispersion of a series of results around their mean, equal to the positive square root of
the variance and estimated by the positive square root of the mean square
[SOURCE: ISO 4259-1:2017, 3.21]
3.3.3
coefficient of variation
ratio of the standard deviation (3.3.2) to the arithmetic mean (3.3.1)
[SOURCE: ISO 27448:2009, 3.11]
3.3.4
relative standard error
standard error (SE ) divided by the mean (x ) and expressed as a percentage
x
3.3.5
analysis of variance
ANOVA
technique which subdivides the total variation of a response variable into components associated with
defined sources of variation
3.3.6
p-value
probability of observing the observed test statistic value or any other value at least as unfavorable to
the null hypothesis
Note 1 to entry: If the null hypothesis were true and if the experiment were repeated many times, a p-value is the
probability that a value at least as extreme as the computed test statistic would be observed.
Note 2 to entry: In hypothesis testing, a statement claiming that the null parameter is the true parameter is
called the null hypothesis. The purpose of a hypothesis test is to determine whether the data provide evidence
against the null hypothesis. When a statistic is obtained that is very different from the null parameter, the null
hypothesis can be rejected. An alternative, or research hypothesis, is a hypothesis that states that the true
parameter is not (or is less than or is greater than) the null parameter; it is the hypothesis that corresponds
to the research question. The goal of a hypothesis test is to reject the null hypothesis in favour of the research
hypothesis.
[SOURCE: ISO/TR 14468:2010, 3.13, modified — Note 1 to entry has been modified and Note 2 to entry
has been added.]
3.3.7
residual deviation
difference between the observed value of the response variable and the estimated value of the response
variable
3.3.8
residual standard deviation
scatter of the information values about the calculated regression line
Note 1 to entry: It is a figure of merit, describing the precision (3.5.3) of the calibration.
Note 2 to entry: For this document, the standard deviation (3.3.2) of the method means the standard of deviation
of the calibration procedure.
[SOURCE: ISO 8466-1:1990, 2.5, modified — the symbol has been deleted and the entire entry has been
editorially revised.]
© ISO 2021 – All rights reserved 5
---------------------- Page: 11 ----------------------
ISO 19749:2021(E)
3.3.9
quantile plot
graphical method of comparing two distributions where the quantiles of the empirical (data)
distribution are plotted on the y-axis while the quantiles of the theoretical (reference) distribution with
the same mean and variance as the empirical distribution are plotted on the x-axis
Note 1 to entry: The quantile-quantile (q-q) plot is a probability plot, a graphical technique for determining if two
data sets come from populations with a common distribution. A q-q plot is a plot of the quantiles of the first data
set against the quantiles of the second data set. See ISO/TS 80004-6.
3.4 Core terms: measurands and descriptors
3.4.1
measurand
quantity intended to be measured
[SOURCE: ISO/IEC Guide 98-4:2012, 3.2.4]
3.4.2
Feret diameter
distance between two parallel lines which are tangent to the perimeter (3.4.5) of a particle (3.1.3)
[SOURCE: ISO 10788:2014, 2.1.4, modified — Note 1 to entry has been deleted.]
3.4.3
maximum Feret diameter
maximum length of an object whatever its orientation
[SOURCE: ISO/TR 945-2:2011, 2.1, modified — the word "Féret" in the term has been changed to "Feret"
and Note 1 to entry has been deleted.]
3.4.4
minimum Feret diameter
minimum length of an object whatever its orientation
Note 1 to entry: The Feret diameter (3.4.2) or Feret's diameter is a measure of an object size along a specified
direction; it is applied to projections of a three-dimensional object on a two-dimensional plane, see Figure 2. It is
also called the caliper diameter.
Key
1 vertical Feret diameter 3 breadth
2 horizontal Feret diameter 4 length
Figure 2 — Horizontal Feret diameter (88 nm) and vertical Feret diamet
...
NORME ISO
INTERNATIONALE 19749
Première édition
2021-07
Nanotechnologies — Détermination
de la distribution de taille et de
forme des particules par microscopie
électronique à balayage
Nanotechnologies — Measurements of particle size and shape
distributions by scanning electron microscopy
Numéro de référence
ISO 19749:2021(F)
©
ISO 2021
---------------------- Page: 1 ----------------------
ISO 19749:2021(F)
DOCUMENT PROTÉGÉ PAR COPYRIGHT
© ISO 2021
Tous droits réservés. Sauf prescription différente ou nécessité dans le contexte de sa mise en œuvre, aucune partie de cette
publication ne peut être reproduite ni utilisée sous quelque forme que ce soit et par aucun procédé, électronique ou mécanique,
y compris la photocopie, ou la diffusion sur l’internet ou sur un intranet, sans autorisation écrite préalable. Une autorisation peut
être demandée à l’ISO à l’adresse ci-après ou au comité membre de l’ISO dans le pays du demandeur.
ISO copyright office
Case postale 401 • Ch. de Blandonnet 8
CH-1214 Vernier, Genève
Tél.: +41 22 749 01 11
E-mail: copyright@iso.org
Web: www.iso.org
Publié en Suisse
ii © ISO 2021 – Tous droits réservés
---------------------- Page: 2 ----------------------
ISO 19749:2021(F)
Sommaire Page
Avant-propos .v
Introduction .vi
1 Domaine d’application . 1
2 Références normatives . 1
3 Termes et définitions . 2
4 Principes généraux .12
4.1 Imagerie MEB .12
4.2 Mesurages de taille d’une particule fondés sur une image MEB.12
4.3 Mesurages de forme d’une particule fondés sur une image MEB .14
5 Préparation des échantillons .14
5.1 Renseignements fondamentaux concernant la préparation des échantillons .14
5.2 Recommandations générales.14
5.3 Garantie d’une bonne préparation d’échantillon de poudre ou de matériaux bruts
dispersés dans un liquide .15
5.3.1 Poudres .15
5.3.2 Dispersions de nanoparticules dans des liquides .15
5.4 Garantie d’une dispersion représentative .15
5.5 Dépôt de nanoparticules sur un substrat .16
5.5.1 Généralités .16
5.5.2 Dépôt de nanoparticules sur des plaquettes de silicium ou d’autres matériaux .16
5.5.3 Dépôts de nanoparticules sur des grilles MET .18
5.6 Nombre d’échantillons à préparer .19
5.7 Nombre de particules à mesurer pour en déterminer la taille .19
5.8 Nombre de particules à mesurer pour en déterminer la forme .20
6 Qualification du MEB pour les mesurages des nanoparticules.20
7 Acquisition d’images .21
7.1 Généralités .21
7.2 Réglage d’une résolution et d’un grandissement corrects de l’image .25
8 Analyse de particules .26
8.1 Renseignements fondamentaux concernant l’analyse de particules .26
8.2 Analyse de particules individuelles .27
8.3 Analyse de particules automatisée .27
8.4 Exemple de mode opératoire d’analyse automatisée des particules .28
9 Analyse des données.29
9.1 Généralités .29
9.2 Tri des données brutes: détections des particules en contact, des artefacts et
des contaminants .29
9.3 Ajustement des modèles aux données .30
9.4 Évaluation de l’incertitude de mesure .30
9.4.1 Généralités .30
9.4.2 Exemple: Incertitude de mesure pour les mesurages de taille de particules .30
9.4.3 Analyse à deux variables .31
10 Rapport de résultats .31
Annexe A (normative) Qualification du MEB pour les mesurages des nanoparticules .33
Annexe B (informative) Préparation d’échantillons transversaux de dioxyde de titane .38
Annexe C (informative) Étude de cas portant sur des nanoparticules bien dispersées
de dioxyde de silicium de 60 nm .40
© ISO 2021 – Tous droits réservés iii
---------------------- Page: 3 ----------------------
ISO 19749:2021(F)
Annexe D (informative) Étude de cas portant sur des nanoparticules de dioxyde de titane
de 40 nm .50
Annexe E (informative) Exemple d’extraction de résultats de taille des particules issus
de mesurages de nanoparticules par MEB à l’aide d’ImageJ .59
Annexe F (informative) Effets de certains paramètres d’acquisition d’images et
des méthodes de seuillage sur les mesurages de taille de particules par MEB .61
Annexe G (informative) Exemple de rapport de résultats de mesurages de nanoparticules
par MEB .66
Bibliographie .76
iv © ISO 2021 – Tous droits réservés
---------------------- Page: 4 ----------------------
ISO 19749:2021(F)
Avant-propos
L’ISO (Organisation internationale de normalisation) est une fédération mondiale d’organismes
nationaux de normalisation (comités membres de l’ISO). L’élaboration des Normes internationales est
en général confiée aux comités techniques de l’ISO. Chaque comité membre intéressé par une étude
a le droit de faire partie du comité technique créé à cet effet. Les organisations internationales,
gouvernementales et non gouvernementales, en liaison avec l’ISO participent également aux travaux.
L’ISO collabore étroitement avec la Commission électrotechnique internationale (IEC) en ce qui
concerne la normalisation électrotechnique.
Les procédures utilisées pour élaborer le présent document et celles destinées à sa mise à jour sont
décrites dans les Directives ISO/IEC, Partie 1. Il convient, en particulier de prendre note des différents
critères d’approbation requis pour les différents types de documents ISO. Le présent document a été
rédigé conformément aux règles de rédaction données dans les Directives ISO/IEC, Partie 2 (voir www
.iso .org/ directives).
L’attention est attirée sur le fait que certains des éléments du présent document peuvent faire l’objet de
droits de propriété intellectuelle ou de droits analogues. L’ISO ne saurait être tenue pour responsable
de ne pas avoir identifié de tels droits de propriété et averti de leur existence. Les détails concernant
les références aux droits de propriété intellectuelle ou autres droits analogues identifiés lors de
l’élaboration du document sont indiqués dans l’Introduction et/ou dans la liste des déclarations de
brevets reçues par l’ISO (voir www .iso .org/ brevets).
Les appellations commerciales éventuellement mentionnées dans le présent document sont données
pour information, par souci de commodité, à l’intention des utilisateurs et ne sauraient constituer un
engagement.
Pour une explication de la nature volontaire des normes, la signification des termes et expressions
spécifiques de l’ISO liés à l’évaluation de la conformité, ou pour toute information au sujet de l’adhésion
de l’ISO aux principes de l’Organisation mondiale du commerce (OMC) concernant les obstacles
techniques au commerce (OTC), voir le lien suivant: www .iso .org/ iso/ fr/ avant -propos.
Le présent document a été préparé par le Comité technique ISO/TC 229, Nanotechnologies.
Il convient que l’utilisateur adresse tout retour d’information ou toute question concernant le présent
document à l’organisme national de normalisation de son pays. Une liste exhaustive desdits organismes
se trouve à l’adresse www .iso .org/ fr/ members .html.
© ISO 2021 – Tous droits réservés v
---------------------- Page: 5 ----------------------
ISO 19749:2021(F)
Introduction
Le présent document fournit des recommandations pour mesurer et établir des rapports sur les
distributions de taille et de forme des particules à l’échelle nanométrique, en utilisant les images
obtenues par un microscope électronique à balayage (MEB). Le présent document s’applique également
au mesurage par MEB des particules de plus grande taille. Les nanoparticules sont des objets
tridimensionnels (3D), mais l’image MEB n’est qu’une représentation bidimensionnelle (2D) de la forme
3D observée avec un angle de vue particulier. L’image MEB fournit de précieuses informations sur la
taille et la forme des particules. Bien que l’image MEB contienne une certaine quantité d’informations
3D, pour des raisons de simplicité, le présent document ne traite pas de la reconstruction des
informations 3D. Une caractérisation tridimensionnelle rigoureuse des nanoparticules inclurait la
taille, la forme, la structure de surface (par exemple, la texture), la composition de la surface et des
matériaux internes ainsi que leurs emplacements dans le volume 3D étudié. Le présent document
porte sur deux attributs de morphologie, à savoir la taille et la forme, pour des nano-objets discrets
et agrégés (matériaux ayant au moins une dimension à l’échelle nanométrique, c’est-à-dire entre 1 nm
et 100 nm). Une préparation appropriée de l’échantillon est essentielle pour obtenir des images de
qualité avec un microscope électronique et les techniques préférées varient souvent selon le matériau
de l’échantillon. Il est tout aussi important de s’assurer que le MEB lui-même est capable d’effectuer les
mesurages avec l’incertitude exigée. Les recommandations habituelles suggèrent de mesurer un grand
nombre de particules, c’est-à-dire plusieurs centaines ou plusieurs milliers, afin d’obtenir des résultats
statistiquement fiables pour les distributions de taille et de forme. Le nombre réel de nano-objets à
mesurer dépend de l’échantillon, de l’incertitude exigée et des performances du MEB. L’évaluation
statistique des données et l’évaluation de l’incertitude des mesurandes font partie intégrante des modes
opératoires de mesure et de rédaction de rapports.
Le présent document contient des articles sur les modes opératoires de mesure, l’analyse des particules
et des données et la rédaction de rapports. Les annexes fournissent des exemples spécifiques de
mesurages ainsi que des recommandations concernant la qualification du MEB pour des mesurages
quantitatifs fiables. L’automatisation de l’acquisition d’images et de l’analyse des données peut réduire
le coût et améliorer la qualité des résultats. Les mesurages d’échantillons de nanoparticules isolées sont
généralement plus faciles à effectuer avec des systèmes automatisés d’acquisition d’images et d’analyse
de particules. Les mesurages de nanoparticules isolées complexes et d’agrégats ou d’agglomérats de
nanoparticules peuvent nécessiter une acquisition d’images et une analyse assistées d’un opérateur.
L’évaluation de la forme des particules est facilitée par un grand nombre de solutions logicielles
d’analyse pertinentes qui permettent également la sélection automatique de divers attributs de forme.
vi © ISO 2021 – Tous droits réservés
---------------------- Page: 6 ----------------------
NORME INTERNATIONALE ISO 19749:2021(F)
Nanotechnologies — Détermination de la distribution
de taille et de forme des particules par microscopie
électronique à balayage
1 Domaine d’application
Le présent document spécifie des méthodes permettant de déterminer les distributions de taille et
de forme des nanoparticules, par l’acquisition et l’évaluation d’images obtenues avec un microscope
électronique à balayage, puis l’obtention de résultats exacts et la rédaction de rapports.
NOTE 1 Le présent document s’applique aux particules dont la limite de taille inférieure dépend de l’incertitude
exigée et des performances appropriées du MEB, après démonstration de sa conformité aux exigences décrites
dans le présent document.
NOTE 2 Le présent document s’applique également aux mesurages par MEB de taille et de forme des particules
de taille supérieure à l’échelle nanométrique.
2 Références normatives
Les documents suivants sont cités dans le texte de sorte qu’ils constituent, pour tout ou partie de leur
contenu, des exigences du présent document. Pour les références datées, seule l’édition citée s’applique.
Pour les références non datées, la dernière édition du document de référence s’applique (y compris les
éventuels amendements).
Guide ISO/IEC 99, Vocabulaire international de métrologie — Concepts fondamentaux et généraux et
termes associés (VIM)
ISO 9276-1, Représentation de données obtenues par analyse granulométrique — Partie 1: Représentation
graphique
ISO 9276-2, Représentation de données obtenues par analyse granulométrique — Partie 2: Calcul des
tailles/diamètres moyens des particules et des moments à partir de distributions granulométriques
ISO 9276-3, Représentation de données obtenues par analyse granulométrique — Partie 3: Ajustement
d'une courbe expérimentale à un modèle de référence
ISO 9276-5, Représentation de données obtenues par analyse granulométrique — Partie 5: Méthodes de
calcul relatif à l'analyse granulométrique à l'aide de la distribution de probabilité logarithmique normale
ISO 9276-6, Représentation de données obtenues par analyse granulométrique — Partie 6: Description et
représentation quantitative de la forme et de la morphologie des particules
ISO 13322-1, Analyse granulométrique — Méthodes par analyse d'images — Partie 1: Méthodes par analyse
d'images statiques
ISO 16700,Analyse par microfaisceaux — Microscopie électronique à balayage — Lignes directrices pour
l'étalonnage du grandissement d'image
ISO/IEC 17025,Exigences générales concernant la compétence des laboratoires d'étalonnages et d'essais
ISO/TS 24597:2011, Analyse par microfaisceaux — Microscopie électronique à balayage — Méthodes
d'évaluation de la netteté d'image
ISO 26824, Caractérisation des particules dans les systèmes particulaires — Vocabulaire
ISO/TS 80004-1, Nanotechnologies — Vocabulaire — Partie 1: Termes "coeur"
© ISO 2021 – Tous droits réservés 1
---------------------- Page: 7 ----------------------
ISO 19749:2021(F)
ISO/TS 80004-2, Nanotechnologies — Vocabulaire — Partie 2: Nano-objets
ISO/TS 80004-3, Nanotechnologies — Vocabulaire — Partie 3: Nano-objets carbonés
ISO/TS 80004-4, Nanotechnologies — Vocabulaire — Partie 4: Matériaux nanostructurés
ISO/TS 80004-6, Nanotechnologies — Vocabulaire — Partie 6: Caractérisation des nano-objets
3 Termes et définitions
Pour les besoins du présent document, les termes et définitions de le Guide ISO/IEC 99, l’ISO 9276-6,
l’ISO 26824, l’ISO/TS 80004-1, l’ISO/TS 80004-2, l’ISO/TS 80004-3, l’ISO/TS 80004-4, l’ISO/TS 80004-6,
ainsi que les suivants, s’appliquent.
L’ISO et l’IEC tiennent à jour des bases de données terminologiques destinées à être utilisées en
normalisation, consultables aux adresses suivantes:
— ISO Online browsing platform: disponible à l’adresse https:// www .iso .org/ obp;
— IEC Electropedia: disponible à l’adresse https:// www .electropedia .org/ .
3.1 Termes généraux
3.1.1
échelle nanométrique
échelle de longueur s’étendant approximativement de 1 nm à 100 nm
Note 1 à l'article: Les propriétés qui ne constituent pas des extrapolations par rapport à des dimensions plus
grandes sont principalement manifestes dans cette échelle de longueur.
[SOURCE: ISO/TS 80004-1:2015, 2.1]
3.1.2
nano-objet
portion discrète de matériau dont une, deux ou les trois dimensions externes sont à l’échelle
nanométrique (3.1.1)
[SOURCE: ISO/TS 80004-1:2015, 2.5, modifiée — La Note 1 à l’article et la source ont été supprimées.]
3.1.3
particule
élément de matière isolé possédant des limites physiques définies
[SOURCE: ISO/TR 16197:2014, 3.10, modifiée — Les Notes 1, 2 et 3 à l’article et la source ont été
supprimées.]
3.1.4
particule primaire
particule source initiale (3.1.3) des agglomérats (3.1.5) ou des agrégats (3.1.6) ou de mélanges de ceux-
ci
[SOURCE: ISO 26824:2013, 1.4, modifiée — Les Notes 1, 2 et 3 à l’article ont été supprimées.]
3.1.5
agglomérat
ensemble de particules (3.1.3) faiblement ou moyennement liées, dont l’aire de la surface externe
résultante est similaire à la somme des aires de surface de chacun des composants
Note 1 à l'article: Le terme «agglomérat» découle du terme latin «agglomerare» qui signifie «former une boule».
Note 2 à l'article: Les forces assurant la cohésion d’un agglomérat sont faibles, par exemple des forces de Van der
Waals ou des forces résultant d’un simple enchevêtrement physique.
2 © ISO 2021 – Tous droits réservés
---------------------- Page: 8 ----------------------
ISO 19749:2021(F)
Note 3 à l'article: Les agglomérats sont également appelés particules secondaires et les particules sources
initiales sont appelées particules primaires (3.1.4).
[SOURCE: ISO 26824:2013, 1.2, modifiée — La Note 1 à l’article a été ajoutée.]
3.1.6
agrégat
particule (3.1.3) composée de particules fortement liées ou fusionnées, dont l’aire de la surface externe
résultante est significativement plus petite que la somme des aires de surface de chacun des composants
Note 1 à l'article: Les forces assurant la cohésion d’un agrégat sont puissantes, par exemple des liaisons covalentes,
ou des forces résultant d’un frittage ou d’un enchevêtrement physique complexe, ou sinon d’anciennes particules
primaires combinées (3.1.4).
Note 2 à l'article: Le terme «agrégat» découle du terme latin «aggregat» qui signifie «rassemblé».
Note 3 à l'article: La Figure 1 montre des exemples de particules individuelles, d’agglomérats (3.1.5) de particules
et d’agrégats de particules.
NOTE Les images sont des vues en projection obtenues à partir de certains angles d’objets 3D. La taille
observable des particules peut varier sensiblement selon l’angle de vue.
Figure 1 — Images MEB de particules d’or (gauche), d’agrégats de noir de carbone (milieu) et
d’agglomérats de corindon (droite)
[SOURCE: ISO 26824:2013, 1.3, modifiée — Les Notes 2 et 3 à l’article ont été ajoutées.]
3.1.7
nanoparticule
nano-objet (3.1.2) dont toutes les dimensions externes sont à l’échelle nanométrique (3.1.1) et dont les
longueurs du plus grand et du plus petit axes ne diffèrent pas de façon significative
[SOURCE: ISO/TS 80004-2:2015, 4.4, modifiée — La Note 1 à l’article a été supprimée.]
3.1.8
taille d’une particule
x
dimension d’une particule (3.1.3) déterminée par une méthode de mesure spécifiée dans des conditions
de mesure spécifiées
Note 1 à l'article: Différentes méthodes d’analyse sont fondées sur le mesurage de différentes propriétés
physiques. Indépendamment de la propriété de particule réellement mesurée, la taille de la particule peut être
consignée comme une dimension linéaire, une surface ou un volume.
Note 2 à l'article: Le symbole x est utilisé pour indiquer la taille linéaire d’une particule (3.1.3). Cependant, il est
reconnu que le symbole d est également couramment utilisé. Le symbole x peut donc être remplacé par d.
© ISO 2021 – Tous droits réservés 3
---------------------- Page: 9 ----------------------
ISO 19749:2021(F)
3.1.9
distribution granulométrique
distribution de le quantité de particules (3.1.3) en fonction de leur taille (3.1.8)
[SOURCE: ISO/TS 80004-6:2021, 4.1.2, modifiée — Les Notes 1 et 2 à l’article ont été supprimée.]
3.1.10
forme d’une particule
forme géométrique externe d’une particule (3.1.3)
Note 1 à l'article: La description de la forme nécessite deux descripteurs scalaires: la longueur et la largeur.
[SOURCE: ISO/TS 80004-6:2013, 3.1.3, modifiée — La Note 1 à l’article a été ajoutée.]
3.1.11
échantillon pour analyse
prise de matériau, issue de l’échantillon d’origine ou d’un échantillon composite, au moyen d’une
méthode appropriée de traitement préalable des échantillons, et ayant la taille (volume/masse)
nécessaire pour les essais ou l’analyse souhaités
Note 1 à l'article: En chimie analytique l’échantillon est une prise de matériau réalisée dans un volume de
matériau plus important. Le terme doit être défini, par exemple échantillon global, échantillon représentatif,
échantillon primaire, échantillon massif, échantillon pour essai. Le terme «échantillon» implique l’existence
d’une erreur d’échantillonnage, c’est-à-dire que les résultats obtenus à partir des prises réalisées ne sont que des
estimations de la concentration d’un constituant ou de la quantité d’une propriété présente dans le matériau de
base. Si l’erreur d’échantillonnage est nulle ou négligeable, la prise effectuée est une prise d’essai, une aliquote
ou un spécimen. Le terme «spécimen» est utilisé pour indiquer une prise effectuée dans des conditions où la
variabilité d’échantillonnage ne peut pas être évaluée (généralement en raison de la variation de la population),
et où elle est considérée nulle par commodité. Il convient d’établir la façon dont l’échantillon est sélectionné dans
un plan d’échantillonnage.
[SOURCE: ISO 11074:2015, 4.1.3, modifiée — La Note 1 à l’article a été ajoutée.]
3.2 Termes «cœur»: analyse d’image
3.2.1
image binaire
image numérisée constituée d’une matrice de pixels (3.2.2), possédant chacun une valeur 0 ou 1, dont
les valeurs sont normalement représentées par des régions sombres et claires sur l’écran d’affichage ou
par l’utilisation de deux couleurs distinctes
[SOURCE: ISO 13322-1:2014, 3.1.2]
3.2.2
pixel
plus petit élément d’une image pouvant être traité de façon unique, qui est défini par ses coordonnées
spatiales et codé avec des valeurs de couleurs
[SOURCE: ISO 12640-2:2004, 3.6, modifiée — La Note 1 à l’article a été supprimée.]
3.2.3
résolution de pixels
nombre de pixels (3.2.2) d’imagerie par unité de distance d’un détecteur
Note 1 à l'article: L’unité type est souvent exprimée en points par pouce (ppp).
[SOURCE: ISO 29301:2017, 3.24, modifiée]
4 © ISO 2021 – Tous droits réservés
---------------------- Page: 10 ----------------------
ISO 19749:2021(F)
3.3 Termes «cœur»: symboles statistiques et définitions
3.3.1
moyenne arithmétique
somme des valeurs divisée par le nombre de valeurs
Note 1 à l'article: Voir l’ISO 9276-1:1998 pour les autres mesures de quantité et types de distributions.
3.3.2
écart-type
mesure de la dispersion d’une série de résultats autour de leur moyenne, égale à la racine carrée positive
de la variance et estimée par la racine carrée positive du carré moyen
[SOURCE: ISO 4259-1:2017, 3.21]
3.3.3
coefficient de variation
rapport de l’écart-type (3.3.2) à la moyenne arithmétique (3.3.1)
[SOURCE: ISO 27448:2009, 3.11]
3.3.4
erreur-type relative (RSE)
erreur-type (SE ) divisée par la moyenne (x ) et exprimée en pourcentage
x
3.3.5
analyse de variance
ANOVA
méthode consistant à séparer la variation totale d’une variable de réponse en composantes associées à
des sources spécifiques de variation
3.3.6
valeur p
probabilité d’obtenir la valeur de la statistique de test observée ou toute autre valeur défavorable à
l’hypothèse nulle
Note 1 à l'article: Si l’hypothèse nulle était vraie et si l’expérience a été répétée un grand nombre de fois, une
valeur p est la probabilité qu’une valeur au moins aussi extrême que la statistique d’essai calculée soit observée.
Note 2 à l'article: Lors d’un test d’hypothèse, une expression affirmant que le paramètre nul est le paramètre
vrai est appelée hypothèse nulle. L’objectif d’un test d’hypothèse est de déterminer si les données fournissent
des preuves contre l’hypothèse nulle. Lorsqu’une statistique obtenue est très différente du paramètre nul,
l’hypothèse nulle peut être rejetée. Une hypothèse alternative ou de recherche est une hypothèse indiquant que
le paramètre vrai n’est pas (ou est inférieur ou supérieur) le paramètre nul. Il s’agit de l’hypothèse correspondant
à la question de recherche. Le but d’un test d’hypothèse est de rejeter l’hypothèse nulle en faveur de l’hypothèse
de recherche.
[SOURCE: ISO/TR 14468:2010, 3.13, modifiée — La Note 1 à l’article a été modifié et la Note 2 à l’article
a été ajoutée.]
3.3.7
écart résiduel
différence entre la valeur observée de la variable de réponse et sa valeur estimée
3.3.8
écart-type résiduel
dispersion des valeurs d’information autour de la ligne de régression calculée
Note 1 à l'article: C’est un indice de performance qui décrit la fidélité (3.5.3) de l’étalonnage.
Note 2 à l'article: Dans le présent document, l’écart-type (3.3.2) de la méthode est l’écart-type de la performance
de l’étalonnage.
© ISO 2021 – Tous droits réservés 5
---------------------- Page: 11 ----------------------
ISO 19749:2021(F)
[SOURCE: ISO 8466-1:1990, 2.5, modifiée — Le symbole a été supprimé et une révision éditori
...
INTERNATIONAL ISO
STANDARD 19749
First edition
2021-05
Nanotechnologies — Measurements of
particle size and shape distributions
by scanning electron microscopy
Nanotechnologies — Détermination de la distribution de taille et de
forme des particules par microscopie électronique à balayage
PROOF/ÉPREUVE
Reference number
ISO 19749:2021(E)
©
ISO 2021
---------------------- Page: 1 ----------------------
ISO 19749:2021(E)
COPYRIGHT PROTECTED DOCUMENT
© ISO 2021
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
be reproduced or utilized otherwise in any form or by any means, electronic or mechanical, including photocopying, or posting
on the internet or an intranet, without prior written permission. Permission can be requested from either ISO at the address
below or ISO’s member body in the country of the requester.
ISO copyright office
CP 401 • Ch. de Blandonnet 8
CH-1214 Vernier, Geneva
Phone: +41 22 749 01 11
Email: copyright@iso.org
Website: www.iso.org
Published in Switzerland
ii PROOF/ÉPREUVE © ISO 2021 – All rights reserved
---------------------- Page: 2 ----------------------
ISO 19749:2021(E)
Contents Page
Foreword .v
Introduction .vi
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 2
3.1 General terms . 2
3.2 Core terms: image analysis . 4
3.3 Core terms: statistical symbols and definitions . 4
3.4 Core terms: measurands and descriptors . 6
3.5 Core terms: metrology . 8
3.6 Core terms: scanning electron microscopy .10
4 General principles .11
4.1 SEM imaging .11
4.2 SEM image-based particle size measurements .12
4.3 SEM image-based particle shape measurements .13
5 Sample preparation .13
5.1 Sample preparation fundamental information .13
5.2 General recommendations.14
5.3 Ensuring good sampling of powder or dispersion-in-liquid raw materials .14
5.3.1 Powders .14
5.3.2 Nanoparticle dispersions in liquids .15
5.4 Ensuring representative dispersion .15
5.5 Nanoparticle deposition on a substrate .15
5.5.1 General.15
5.5.2 Nanoparticle deposition on wafers and chips of silicon or other materials .16
5.5.3 Nanoparticle deposition on TEM grids .17
5.6 Number of samples to be prepared .18
5.7 Number of particles to be measured for particle size determination .18
5.8 Number of particles to be measured for particle shape determination .19
6 Qualification of the SEM for nanoparticle measurements .19
7 Image acquisition .19
7.1 General .19
7.2 Setting suitable image magnification and pixel resolution .23
8 Particle analysis .24
8.1 Particle analysis fundamental information .24
8.2 Individual particle analysis .25
8.3 Automated particle analysis .25
8.4 Automated particle analysis procedure example .26
9 Data analysis .26
9.1 General .26
9.2 Raw data screening: detecting touching particles, artefacts and contaminants .27
9.3 Fitting models to data .27
9.4 Assessment of measur ement uncertainty .27
9.4.1 General.27
9.4.2 Example: measurement uncertainty for particle size measurements .28
9.4.3 Bivariate analysis .28
10 Reporting the results .29
Annex A (normative) Qualification of the SEM for nanoparticle measurements .30
Annex B (informative) Cross-sectional titanium dioxide samples preparation .35
© ISO 2021 – All rights reserved PROOF/ÉPREUVE iii
---------------------- Page: 3 ----------------------
ISO 19749:2021(E)
Annex C (informative) Case study on well-dispersed 60 nm size silicon dioxide nanoparticles .37
Annex D (informative) Case study on 40 nm size titanium dioxide nanoparticles .45
Annex E (informative) Example for extracting particle size results of SEM-based
nanoparticle measurements using ImageJ .54
Annex F (informative) Effects of some image acquisition parameters and thresholding
methods on SEM particle size measurements .56
Annex G (informative) Example for reporting results of SEM-based nanoparticle measurements 60
Bibliography .70
iv PROOF/ÉPREUVE © ISO 2021 – All rights reserved
---------------------- Page: 4 ----------------------
ISO 19749:2021(E)
Foreword
ISO (the International Organization for Standardization) is a worldwide federation of national standards
bodies (ISO member bodies). The work of preparing International Standards is normally carried out
through ISO technical committees. Each member body interested in a subject for which a technical
committee has been established has the right to be represented on that committee. International
organizations, governmental and non-governmental, in liaison with ISO, also take part in the work.
ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of
electrotechnical standardization.
The procedures used to develop this document and those intended for its further maintenance are
described in the ISO/IEC Directives, Part 1. In particular, the different approval criteria needed for the
different types of ISO documents should be noted. This document was drafted in accordance with the
editorial rules of the ISO/IEC Directives, Part 2 (see www .iso .org/ directives).
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. ISO shall not be held responsible for identifying any or all such patent rights. Details of
any patent rights identified during the development of the document will be in the Introduction and/or
on the ISO list of patent declarations received (see www .iso .org/ patents).
Any trade name used in this document is information given for the convenience of users and does not
constitute an endorsement.
For an explanation of the voluntary nature of standards, the meaning of ISO specific terms and
expressions related to conformity assessment, as well as information about ISO's adherence to the
World Trade Organization (WTO) principles in the Technical Barriers to Trade (TBT), see www .iso .org/
iso/ foreword .html.
This document was prepared jointly by Technical Committee ISO/ TC 229, Nanotechnologies, and
Technical Committee IEC/TC 113, Nanotechnology for electrotechnical products and systems.
Any feedback or questions on this document should be directed to the user’s national standards body. A
complete listing of these bodies can be found at www .iso .org/ members .html.
© ISO 2021 – All rights reserved PROOF/ÉPREUVE v
---------------------- Page: 5 ----------------------
ISO 19749:2021(E)
Introduction
This document provides guidance for measuring and reporting the size and shape distributions of
nanometer-scale particles using images acquired by the scanning electron microscope (SEM). This
document applies to the SEM-based measurement of larger particles also. Nanoparticles are three-
dimensional (3D) objects, but the SEM image is only a two-dimensional (2D) representation of the 3D
shape from a certain viewing angle. The SEM image carries valuable information about the size and
shape of particles. While the SEM image does contain a certain amount of 3D information, for sake of
simplicity, this document does not deal with reconstructing 3D information. Rigorous three-dimensional
characterization of nanoparticles would include size, shape, surface structure (e.g. texture), surface
and internal material composition, and their locations in the investigated 3D volume. This document
deals with two attributes of morphology, size and shape, for discrete and aggregated nano-objects
(materials with at least one dimension in the nanometer-scale, i.e. within 1 nm to 100 nm). Suitable
sample preparation is essential to obtaining high-quality electron microscope images and preferred
techniques often vary with the sample material. It is equally important to make sure that the SEM itself
is suitable to carry out the measurements with the required uncertainty. Typical guidance suggests that
a large number, several hundreds or thousands of particles need to be measured for statistically sound
size and shape distribution results. The actual number of nano-objects needed to be measured depends
on the sample, the required uncertainty and on the performance of the SEM. Statistical evaluation of
the data and the evaluation of uncertainty of the measurands are included as part of the measurement
and reporting procedures.
This document contains measurement procedures, particle and data analysis and reporting clauses. In
the Annexes, there are specific examples for measurements and guidance for the qualification of the
SEM for reliable quantitative measurements. Automation of the image acquisition and data analysis can
reduce cost and improve the quality of the results. Measurements of samples of discrete nanoparticles
are generally easier to carry out with automated image acquisition and particle analysis systems.
Measurements of complex discrete nanoparticles, and aggregates or agglomerates of nanoparticles
may require operator-assisted image acquisition and analysis. Evaluation of particle shape is facilitated
by many pertinent analysis software solutions that allow for automatic selection of various shape
attributes as well.
vi PROOF/ÉPREUVE © ISO 2021 – All rights reserved
---------------------- Page: 6 ----------------------
INTERNATIONAL STANDARD ISO 19749:2021(E)
Nanotechnologies — Measurements of particle size and
shape distributions by scanning electron microscopy
1 Scope
This document specifies methods of determining nanoparticle size and shape distributions by acquiring
and evaluating scanning electron microscope images and by obtaining and reporting accurate results.
NOTE 1 This document applies to particles with a lower size limit that depends on the required uncertainty
and on the suitable performance of the SEM, which is to be proven first -according to the requirements described
in this document.
NOTE 2 This document applies also to SEM-based size and shape measurements of larger than nanoscale
particles.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
ISO/IEC Guide 99, International vocabulary of metrology — Basic and general concepts and associated
terms (VIM)
ISO 9276-1, Representation of results of particle size analysis — Part 1: Graphical representation
ISO 9276-2, Representation of results of particle size analysis — Part 2: Calculation of average particle
sizes/diameters and moments from particle size distributions
ISO 9276-3, Representation of results of particle size analysis — Part 3: Adjustment of an experimental
curve to a reference model
ISO 9276-5, Representation of results of particle size analysis — Part 5: Methods of calculation relating to
particle size analyses using logarithmic normal probability distribution
ISO 9276-6, Representation of results of particle size analysis — Part 6: Descriptive and quantitative
representation of particle shape and morphology. This document provides a detailed list of other shape
parameters for size, macroshape descriptors, geometrical descriptors, proportion descriptors, and
mesoshape descriptors
ISO 13322-1, Particle size analysis — Image analysis methods — Part 1: Static image analysis methods
ISO 26824, Particle characterization of particulate systems — Vocabulary
ISO 29301, Microbeam analysis — Analytical electron microscopy — Methods for calibrating image
magnification by using reference materials with periodic structures
ISO/TS 80004-1, Nanotechnologies — Vocabulary — Part 1: Core terms
ISO/TS 80004-2, Nanotechnologies — Vocabulary — Part 2: Nano-objects
ISO/TS 80004-3, Nanotechnologies – Vocabulary – Part 3: Carbon nano-objects
ISO/TS 80004-4, Nanotechnologies — Vocabulary — Part 4: Nanostructured materials
ISO/TS 80004-6, Nanotechnologies – Vocabulary – Part 6: Nano-object characterization
© ISO 2021 – All rights reserved PROOF/ÉPREUVE 1
---------------------- Page: 7 ----------------------
ISO 19749:2021(E)
3 Terms and definitions
For the purposes of this document, the terms and definitions given in ISO 9276-6, ISO 26824,
ISO/TS 80004-1, ISO/TS 80004-2, ISO/TS 80004-3, ISO/TS 80004-4, ISO/TS 80004-6, ISO/IEC Guide 99,
and the following apply.
ISO and IEC maintain terminological databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https:// www .iso .org/ obp
— IEC Electropedia: available at http:// www .electropedia .org/
3.1 General terms
3.1.1
nanoscale
length range from approximately 1 nm to 100 nm
Note 1 to entry: Properties that are not extrapolations from larger sizes are predominantly exhibited in this
length range.
[SOURCE: ISO/TS 80004-1:2015, 2.1]
3.1.2
nano-object
discrete piece of material with one, two or three external dimensions in the nanoscale (3.1.1)
[SOURCE: ISO/TS 80004-3:2015, 3.1.3, modified — Note 1 to entry and the source have been deleted.]
3.1.3
particle
minute piece of matter with defined physical boundaries
[SOURCE: ISO/TR 16197:2014, 3.10, modified — Notes 1, 2 and 3 to entry and the source have been
deleted.]
3.1.4
primary particle
original source particle (3.1.3) of agglomerates (3.1.5) or aggregates (3.1.6) or mixtures of the two
[SOURCE: ISO 26824:2013, 1.4, modified — Notes 1, 2 and 3 to entry have been deleted.]
3.1.5
agglomerate
collection of weakly or medium strongly bound particles (3.1.3) where the resulting external surface
area is similar to the sum of the surface areas of the individual components
Note 1 to entry: Agglomerate originates from the Latin “agglomerare” meaning “to form into a ball”.
Note 2 to entry: The forces holding an agglomerate together are weak forces, for example van der Waals forces or
simple physical entanglement.
Note 3 to entry: Agglomerates are also termed secondary particles and the original source particles are termed
primary particles (3.1.4).
[SOURCE: ISO 26824:2013, 1.2, modified — Note 1 to entry has been added.]
2 PROOF/ÉPREUVE © ISO 2021 – All rights reserved
---------------------- Page: 8 ----------------------
ISO 19749:2021(E)
3.1.6
aggregate
particle (3.1.3) comprising strongly bonded or fused particles where the resulting external surface area
is significantly smaller than the sum of surface areas of the individual components
Note 1 to entry: The forces holding an aggregate together are strong forces, for example covalent bonds, or those
resulting from sintering or complex physical entanglement, or otherwise combined former primary particles
(3.1.4).
Note 2 to entry: Aggregate comes from the Latin “aggregat” meaning “herded together”.
Note 3 to entry: Figure 1 shows examples of individual, aggregate and agglomerate (3.1.5) particles.
NOTE The images are projected views from certain angles of the 3D objects. Depending on the viewing
angle, the observable size of particles can vary substantially.
Figure 1 — SEM images of individual gold (left) and carbon black aggregate (middle) and
corundum agglomerate (right) particles
[SOURCE: ISO 26824:2013, 1.3, modified — Notes 2 and 3 to entry have been added.]
3.1.7
nanoparticle
nano-object (3.1.2) with all external dimensions in the nanoscale (3.1.1) where the lengths of the longest
and shortest axes of the nano-object do not differ significantly
[SOURCE: ISO/TS 80004-2:2015, 4.4, modified — Note 1 to entry has been deleted.]
3.1.8
particle size
x
dimension of a particle (3.1.3) determined by a specified measurement method and under specified
measurement conditions
Note 1 to entry: Different methods of analysis are based on the measurement of different physical properties.
Independent of the particle property actually measured, the particle size can be reported as a linear dimension,
an area, or a volume.
Note 2 to entry: The symbol x is used denote linear particle (3.1.3) size. However, it is recognized that the symbol
d is also widely used. Therefore, the symbol x may be replaced by d.
3.1.9
particle size distribution
distribution of particles (3.1.3) as a function of particle size (3.1.8)
[SOURCE: ISO/TS 80004-6:2013, 3.1.2, modified — Note 1 to entry has been deleted.]
© ISO 2021 – All rights reserved PROOF/ÉPREUVE 3
---------------------- Page: 9 ----------------------
ISO 19749:2021(E)
3.1.10
particle shape
external geometric form of a particle (3.1.3)
Note 1 to entry: Shape description requires two scalar descriptors, i.e. length and breadth.
[SOURCE: ISO/TS 80004-6:2013, 3.1.3, modified — Note 1 to entry has been added.]
3.1.11
analytical sample
portion of material, resulting from the original sample or composite sample by means of an appropriate
method of sample pretreatment and having the size (volume/mass) necessary for the desired testing or
analysis
Note 1 to entry: the sample in analytical chemistry is a portion of material selected from a larger quantity of
material. The term needs to be qualified, for example, bulk sample, representative sample, primary sample,
bulked sample, test sample. The term 'sample' implies the existence of a sampling error, i.e. the results obtained
on the portions taken are only estimates of the concentration of a constituent or the quantity of a property present
in the parent material. If there is no or negligible sampling error, the portion removed is a test portion, aliquot
or specimen. The term 'specimen' is used to denote a portion taken under conditions such that the sampling
variability cannot be assessed (usually because the population is changing), and is assumed, for convenience, to
be zero. The manner of selection of the sample should be prescribed in a sampling plan.
[SOURCE: ISO 11074:2015, 4.1.3, modified — Note 1 to entry has been added.]
3.2 Core terms: image analysis
3.2.1
binary image
digitized image consisting of an array of pixels (3.2.2), each of which has a value of 0 or 1, whose values are
normally represented by dark and bright regions on the display screen or by the use of two distinct colors
[SOURCE: ISO 13322-1:2014, 3.1.2]
3.2.2
pixel
smallest element of an image that can be uniquely processed, and is defined by its spatial coordinates
and encoded with colour values
[SOURCE: ISO 12640-2:2004, 3.6, modified — Note 1 to entry has been deleted.]
3.2.3
pixel resolution
number of imaging pixels (3.2.2) per unit distance of the detector
Note 1 to entry: The typical unit is sometimes expressed as dots per inch (dpi).
[SOURCE: ISO 29301:2017, 3.24, modified — the hyphen has been deleted in this term.]
3.3 Core terms: statistical symbols and definitions
3.3.1
arithmetic mean
sum of values divided by the number of values
Note 1 to entry: See ISO 9276-1:1998 for other quantity measures and types of distributions.
4 PROOF/ÉPREUVE © ISO 2021 – All rights reserved
---------------------- Page: 10 ----------------------
ISO 19749:2021(E)
3.3.2
standard deviation
measure of the dispersion of a series of results around their mean, equal to the positive square root of
the variance and estimated by the positive square root of the mean square
[SOURCE: ISO 4259-1:2017, 3.21]
3.3.3
coefficient of variation
ratio of the standard deviation (3.3.2) to the arithmetic mean (3.3.1)
[SOURCE: ISO 27448:2009, 3.11]
3.3.4
relative standard error
standard error (SE ) divided by the mean (x ) and expressed as a percentage
x
3.3.5
analysis of variance
ANOVA
technique which subdivides the total variation of a response variable (2.2) into components associated
with defined sources of variation
3.3.6
p-value
probability of observing the observed test statistic value or any other value at least as unfavorable to
the null hypothesis
Note 1 to entry: If the null hypothesis were true and if the experiment were repeated many times, a p-value is the
probability that a value at least as extreme as the computed test statistic would be observed.
Note 2 to entry: In hypothesis testing, a statement claiming that the null parameter is the true parameter is
called the null hypothesis. The purpose of a hypothesis test is to determine whether the data provide evidence
against the null hypothesis. When a statistic is obtained that is very different from the null parameter, the null
hypothesis can be rejected. An alternative, or research hypothesis, is a hypothesis that states that the true
parameter is not (or is less than or is greater than) the null parameter; it is the hypothesis that corresponds
to the research question. The goal of a hypothesis test is to reject the null hypothesis in favour of the research
hypothesis.
[SOURCE: ISO/TR 14468:2010, 3.13, modified — Notes 1 and 2 to entry have been added.]
3.3.7
residual deviation
difference between the observed value of the response variable and the estimated value of the response
variable
3.3.8
residual standard deviation
scatter of the information values about the calculated regression line. It is a figure of merit, describing
the precision (3.5.3) of the calibrationNote 1 to entry: For this document, the standard deviation (3.3.2)
of the method means the standard of deviation of the calibration procedure.
[SOURCE: ISO 8466-1:1990, 2.5, modified — the symbol has been deleted and the entire entry has been
editorially revised.]
© ISO 2021 – All rights reserved PROOF/ÉPREUVE 5
---------------------- Page: 11 ----------------------
ISO 19749:2021(E)
3.3.9
quantile plot
graphical method of comparing two distributions where the quantiles of the empirical (data)
distribution are plotted on the y-axis while the quantiles of the theoretical (reference) distribution with
the same mean and variance as the empirical distribution are plotted on the x-axis
Note 1 to entry: The quantile-quantile (q-q) plot is a probability plot, a graphical technique for determining if two
data sets come from populations with a common distribution. A q-q plot is a plot of the quantiles of the first data
set against the quantiles of the second data set. See Reference [21].
3.4 Core terms: measurands and descriptors
3.4.1
measurand
quantity intended to be measured
[SOURCE: ISO/IEC Guide 98-4:2012, 3.2.4]
3.4.2
Feret diameter
distance between two parallel lines which are tangent to the perimeter (3.4.5) of a particle (3.1.3)
[SOURCE: ISO 10788:2014, 2.1.4, modified — Note 1 to entry has been deleted.]
3.4.3
maximum Feret diameter
maximum length of an object whatever its orientation
[SOURCE: ISO/TR 945-2:2011, 2.1, modified — the word "Féret" in the term has been changed to "Feret".]
3.4.4
minimum Feret diameter
minimum length of an object whatever its orientation
Note 1 to entry: The Feret diameter (3.4.2) or Feret's diameter is a measure of an object size along a specified
direction; it is applied to projections of a three-dimensional object on a two-dimensional plane, see Figure 2. It is
also called
...
Questions, Comments and Discussion
Ask us and Technical Secretary will try to provide an answer. You can facilitate discussion about the standard in here.