SIST ISO 10312:2019
Ambient air - Determination of asbestos fibres - Direct transfer transmission electron microscopy method
Ambient air - Determination of asbestos fibres - Direct transfer transmission electron microscopy method
This document specifies a reference method using transmission electron microscopy for the determination of airborne asbestos fibres and structures in in a wide range of ambient air situations, including the interior atmospheres of buildings, and for a detailed evaluation for asbestos structures in any atmosphere. The method allows determination of the type(s) of asbestos fibres present and also includes measurement of the lengths, widths and aspect ratios of the asbestos structures. The method cannot discriminate between individual fibres of asbestos and elongate fragments (cleavage fragments and acicular particles) from non-asbestos analogues of the same amphibole mineral[13].
Air ambiant - Dosage des fibres d'amiante - Méthode par microscopie électronique à transmission par transfert direct
Le présent document spécifie une méthode de référence utilisant la microscopie électronique à transmission pour déterminer la concentration en fibres et structures d'amiante en suspension dans l'air dans diverses atmosphères ambiantes, notamment les atmosphères intérieures de bâtiments, et pour évaluer en détail les structures d'amiante dans les atmosphères. Cette méthode permet de déterminer le(s) type(s) de fibres d'amiante présentes et comprend également le mesurage des longueurs, des largeurs et des rapports longueur/largeur des structures d'amiante. Elle ne peut pas faire la différence entre des fibres individuelles d'amiante et des fragments allongés (fragments de clivage et particules aciculaires) d'analogues non asbestiformes du même minéral amphibole[13].
Zunanji zrak - Določevanje azbestnih vlaken - Metoda transmisijske elektronske mikroskopije z neposrednim prenosom
Ta dokument določa referenčno metodo, pri kateri se s prenosno elektronsko mikroskopijo določajo azbestna vlakna in strukture v zraku v najrazličnejših okoliščinah zunanjega zraka, vključno z notranjo atmosfero stavb, ter za podrobno oceno azbestnih struktur v poljubni atmosferi. Metoda omogoča določevanje vrst(-e) prisotnih azbestnih vlaken in vključuje tudi merjenje dolžin, širin in razmerja azbestnih struktur. Z metodo ni mogoče razlikovati med posameznimi vlakni azbesta in razteznimi fragmenti (delci cepitve in acikularnimi delci) iz neazbestnih analogov istega amfibolovega minerala.
General Information
Relations
Overview
SIST ISO 10312:2019 - "Ambient air - Determination of asbestos fibres - Direct transfer transmission electron microscopy method" - is the international reference method adopted by SIST for measuring asbestos fibres in ambient atmospheres. The standard specifies a direct-transfer TEM (transmission electron microscopy) procedure for collecting and analysing airborne asbestos in a wide range of environments, including indoor building atmospheres. It defines how to identify asbestos types and to measure fibre lengths, widths and aspect ratios, and sets out performance, quality control and reporting requirements.
Key technical topics and requirements
- Scope and purpose: Reference method for airborne asbestos fibres and structures in ambient air and interior atmospheres; usable for detailed evaluations.
- Analytical principle: Direct transfer specimen preparation for TEM, combined with particle morphology and chemical analysis (e.g., EDXA), to identify asbestos minerals.
- Measurements reported: Type(s) of asbestos present and dimensional data - lengths, widths and aspect ratios for asbestos structures.
- Limitations: The method cannot reliably discriminate individual asbestos fibres from elongate cleavage fragments or acicular particles from non‑asbestos amphibole analogues.
- Sampling and specimen prep: Procedures for air sampling, filter handling (polycarbonate and cellulose ester filters), carbon coating, plasma ashing and replica techniques are described.
- Equipment and consumables: Transmission electron microscope (TEM), energy dispersive X‑ray analyser (EDXA), plasma asher, vacuum/sputter coaters, sampling pumps and filter cassettes.
- Quality assurance: Blank controls, calibration procedures, structure counting criteria, limits of detection, precision/accuracy assessment and test report content are specified.
- Annexes: Normative and informative annexes cover plasma asher settings, calibration, counting criteria, fibre identification, PCM-equivalent calculations and sampling strategies.
Practical applications and users
SIST ISO 10312:2019 is used by:
- Environmental and analytical laboratories performing asbestos air monitoring
- Occupational and public health agencies assessing exposure in workplaces and public buildings
- Industrial hygienists and environmental consultants conducting indoor air quality surveys and post‑remediation verification
- Regulatory bodies and developers requiring reference‑method asbestos quantification The method supports compliance assessments, risk evaluations and comparative studies (including PCM-equivalent fibre calculations).
Related standards
- This edition replaces the previous SIST/ISO 10312 (1995/1996) edition.
- Annex E provides guidance for calculating PCM-equivalent asbestos fibres, bridging TEM results with phase‑contrast microscopy methods commonly used in regulation.
Keywords: SIST ISO 10312:2019, ambient air, asbestos fibres, transmission electron microscopy, TEM direct transfer, asbestos analysis, air monitoring, ambient atmospheres.
Standards Content (Sample)
SLOVENSKI STANDARD
01-december-2019
Nadomešča:
SIST ISO 10312:1996
Zunanji zrak - Določevanje azbestnih vlaken - Metoda transmisijske elektronske
mikroskopije z neposrednim prenosom
Ambient air - Determination of asbestos fibres - Direct transfer transmission electron
microscopy method
Air ambiant - Dosage des fibres d'amiante - Méthode par microscopie électronique à
transmission par transfert direct
Ta slovenski standard je istoveten z: ISO 10312:2019
ICS:
13.040.20 Kakovost okoljskega zraka Ambient atmospheres
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
INTERNATIONAL ISO
STANDARD 10312
Second edition
2019-10
Ambient air — Determination of
asbestos fibres — Direct transfer
transmission electron microscopy
method
Air ambiant — Dosage des fibres d'amiante — Méthode par
microscopie électronique à transmission par transfert direct
Reference number
©
ISO 2019
© ISO 2019
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
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Published in Switzerland
ii © ISO 2019 – All rights reserved
Contents Page
Foreword .v
Introduction .vi
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Symbols and abbreviated terms . 5
5 Type of sample . 6
6 Range . 6
7 Limit of detection . 6
8 Principle . 7
9 Reagents . 7
10 Apparatus . 8
10.1 Air sampling — Equipment and consumable supplies . 8
10.1.1 Filter cassette . 8
10.1.2 Sampling pump . 8
10.1.3 Stand . 8
10.1.4 Personal sampling . 8
10.1.5 Flowmeter . 8
10.2 Specimen preparation laboratory . 9
10.3 Equipment for analysis . 9
10.3.1 Transmission electron microscope . 9
10.3.2 Energy dispersive X-ray analyser .11
10.3.3 Plasma asher .11
10.3.4 Vacuum coating unit .11
10.3.5 Sputter coater .11
10.3.6 Solvent washer (Jaffe washer) .11
10.3.7 Condensation washer .12
10.3.8 Slide warmer or oven .13
10.3.9 Ultrasonic bath .13
10.3.10 Carbon grating replica.13
10.3.11 Calibration specimen grids for EDXA .13
10.3.12 Carbon rod sharpener .14
10.3.13 Disposable tip micropipettes .14
10.4 Consumable supplies .14
10.4.1 Copper or nickel electron microscope grids .14
10.4.2 Gold or nickel electron microscope grids .14
10.4.3 Carbon rod electrodes .14
10.4.4 Routine electron microscopy tools and supplies .14
10.4.5 Reference asbestos samples . .14
10.4.6 Reference samples of mineral fibres other than asbestos .15
11 Air sample collection .15
12 Procedure for analysis .16
12.1 General .16
12.2 Cleaning of sample cassettes .16
12.3 Direct preparation of TEM specimens from polycarbonate filters .17
12.3.1 Selection of filter area for carbon coating .17
12.3.2 Carbon coating of filter portions .17
12.3.3 Preparation of the Jaffe washer .17
12.3.4 Placing of specimens in the Jaffe washer .17
12.4 Direct preparation of TEM specimens from cellulose ester filters .18
12.4.1 Selection of area of filter for preparation .18
12.4.2 Preparation of solution for collapsing cellulose ester filters .18
12.4.3 Filter collapsing procedure .18
12.4.4 Plasma etching of the filter surfaces .18
12.4.5 Carbon coating .18
12.4.6 Preparation of the Jaffe washer .18
12.4.7 Placing of specimens in the Jaffe washer .18
12.5 Criteria for acceptable TEM specimen grids .19
12.6 Procedure for structure counting by TEM.19
12.6.1 General.19
12.6.2 Measurement of mean opening area .20
12.6.3 TEM alignment and calibration procedures .20
12.6.4 Determination of criterion for termination of TEM examination.20
12.6.5 General procedure for structure counting and size analysis .21
12.6.6 Magnification requirements .23
12.7 Blank and quality control determinations .24
12.8 Calculation of results .24
13 Performance characteristics .25
13.1 General .25
13.2 Interferences and limitations of fibre identification .25
13.3 Precision and accuracy.25
13.3.1 Precision.25
13.3.2 Accuracy .26
13.3.3 Inter-laboratory and intra-laboratory analyses .26
13.4 Limit of detection .26
14 Test report .27
Annex A (normative) Determination of operating conditions for plasma asher .30
Annex B (normative) Calibration procedures.31
Annex C (normative) Structure counting criteria .34
Annex D (normative) Fibre identification procedure .44
Annex E (normative) Determination of the concentration of asbestos fibres and bundles
longer than 5 µm, and PCM equivalent asbestos fibres .61
Annex F (normative) Calculation of results .62
Annex G (informative) Strategies for collection of air samples .68
Annex H (informative) Methods for removal of gypsum fibres .69
Bibliography .70
iv © ISO 2019 – All rights reserved
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 146, Air quality, Subcommittee SC 3,
Ambient atmospheres.
This second edition cancels and replaces the first edition (ISO 10312:1995), which has been technically
revised. The main changes compared to the previous edition are as follows:
— the use of electronic display systems with measurement software is permitted;
— the maximum particulate loading for TEM specimens is increased from 10 % to 25 %;
— a simplified fibre identification procedure for investigation of known sources of the regulated
asbestos varieties and richterite/winchite asbestos is permitted;
— the reporting requirements have been changed to permit reporting of the concentrations of fibres
and bundles longer than 5 µm and/or the concentrations of PCM equivalent fibres without the
requirement to report the concentrations of structures equal to or greater than 0,5 µm;
— there is no requirement to report the 95% confidence intervals of the fibre concentrations.
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.
Introduction
This document is applicable to the determination of airborne asbestos in a wide range of ambient
air situations, including the interior atmospheres of buildings, and for a detailed evaluation of
any atmosphere. Because the best available medical evidence indicates that the numerical fibre
concentration and the fibre sizes are the relevant parameters for evaluation of the inhalation hazards,
a fibre counting technique is the only logical approach. Most fibres in ambient atmospheres are not
asbestos and therefore, there is a requirement for fibres to be identified. Many airborne asbestos
fibres in ambient atmospheres have diameters below the resolution limit of the optical microscope.
This document is based on transmission electron microscopy, which has adequate resolution to allow
detection of small fibres and is currently the only technique capable of unequivocal identification of
the majority of individual fibres of asbestos. Airborne asbestos is often found as a mixture of single
fibres and more complex, aggregated structures which may or may not be also aggregated with other
particles. The fibres found suspended in an ambient atmosphere can often be identified unequivocally,
if a sufficient measurement effort is expended. However, if each fibre were to be identified in this way,
the analysis would become prohibitively expensive. Because of instrumental deficiencies or because
of the nature of the particulate, some fibres cannot be positively identified as asbestos, even though
the measurements all indicate that they could be asbestos. Subjective factors therefore contribute
to this measurement, and consequently a very precise definition of the procedure for identification
and enumeration of, asbestos fibres is required. The method specified in this document is designed
to provide the best description possible of the nature, numerical concentration, and sizes of asbestos-
containing particles found in an air sample. This document requires that a very detailed and logical
procedure be used to reduce the subjective aspects of the measurement. The method of data recording
specified in this document is designed to allow re-evaluation of the structure counting data as new
medical evidence becomes available. All feasible specimen preparation techniques result in some
modification of the airborne particulate. Even the collection of particles from a three-dimensional
airborne dispersion onto a two-dimensional filter surface can be considered a modification of the
particulate, and some of the particles in most samples are modified by the specimen preparation
procedures. However, the procedures specified in this document are designed to minimize the
disturbance of the collected particulate material, and the effect of those disturbances that do occur can
be evaluated.
This document describes the method of analysis for a single air filter. However, one of the largest
potential errors in characterizing asbestos in ambient atmospheres is associated with the variability
between filter samples. For this reason, it is necessary to design a replicate sampling scheme in order to
determine this document’s accuracy and precision.
vi © ISO 2019 – All rights reserved
INTERNATIONAL STANDARD ISO 10312:2019(E)
Ambient air — Determination of asbestos fibres — Direct
transfer transmission electron microscopy method
1 Scope
This document specifies a reference method using transmission electron microscopy for the
determination of airborne asbestos fibres and structures in in a wide range of ambient air situations,
including the interior atmospheres of buildings, and for a detailed evaluation for asbestos structures
in any atmosphere. The method allows determination of the type(s) of asbestos fibres present and also
includes measurement of the lengths, widths and aspect ratios of the asbestos structures. The method
cannot discriminate between individual fibres of asbestos and elongate fragments (cleavage fragments
[13]
and acicular particles) from non-asbestos analogues of the same amphibole mineral .
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 4225, Air quality — General aspects — Vocabulary
3 Terms and definitions
For the purposes of this document, the terms and definitions given in ISO 4225 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
acicular
shape shown by an extremely slender crystal with cross-sectional dimensions, which are small relative
to its length, i.e. needle-like
3.2
amphibole
group of rock-forming ferromagnesium silicate minerals, closely related in crystal form and
composition, and having the nominal formula:
A B C T O (OH,F,Cl)
0-1 2 5 8 22 2
where
A = K, Na;
2+
B = Fe , Mn, Mg, Ca, Na;
3+ 2+
C = Al, Cr, Ti, Fe , Mg, Fe ;
3+
T = Si, Al, Cr, Fe , Ti.
Note 1 to entry: In some varieties of amphibole, these elements can be partially substituted by Li, Pb, or Zn.
Amphibole is characterized by a cross-linked double chain of Si-O tetrahedra with a silicon: oxygen ratio of 4:11,
by columnar or fibrous prismatic crystals and by good prismatic cleavage in two directions parallel to the crystal
faces and intersecting at angles of about 56° and 124°.
3.3
amphibole asbestos
amphibole (3.2) in an asbestiform (3.5) habit
3.4
analytical sensitivity
calculated airborne asbestos structure (3.7) concentration in structures/litre, equivalent to counting of
one asbestos (3.6) structure in the analysis
Note 1 to entry: It is expressed in structures/litre.
Note 2 to entry: This method does not specify a unique analytical sensitivity. The analytical sensitivity is
determined by the needs of the measurement and the conditions found on the prepared sample.
3.5
asbestiform
specific type of mineral fibrosity in which the fibres (3.22) and fibrils possess high tensile strength and
flexibility
3.6
asbestos
group of silicate minerals belonging to the serpentine and amphibole (3.2) groups, which have
crystallized in the asbestiform habit, causing them to be easily separated into long, thin, flexible, strong
fibres (3.22) when crushed or processed
Note 1 to entry: The Chemical Abstracts Service Registry Numbers of the most common asbestos varieties are:
chrysotile (12001-29-5), crocidolite (12001-28-4), grunerite asbestos (Amosite) (12172-73-5), anthophyllite
asbestos (77536-67-5), tremolite asbestos (77536-68-6) and actinolite asbestos (77536-66-4). Other varieties of
[19]
asbestiform amphibole, such as richterite asbestos and winchite asbestos may also be found in some products
such as vermiculite and talc.
3.7
asbestos structure
individual fibre (3.22), or any connected or overlapping grouping of asbestos (3.6) fibres or bundles,
with or without other particles
3.8
aspect ratio
ratio of length to width of a particle
3.9
blank
structure count made on transmission electron microscope specimens prepared from an unused filter,
to determine the background measurement
3.10
camera length
equivalent projection length between the specimen and its electron diffraction pattern, in the absence
of lens action
3.11
chrysotile
fibrous mineral of the serpentine group, which has the nominal composition: Mg Si O (OH)
3 2 5 4
Note 1 to entry: Most natural chrysotile deviates little from this nominal composition. In some varieties of
3+ 3+ 2+ 3+
chrysotile, minor substitution of silicon by Al may occur. Minor substitution of magnesium by Al , Fe , Fe ,
2+ 2+ 2+
Ni , Mn and Co may also be present. Chrysotile is the most prevalent type of asbestos.
2 © ISO 2019 – All rights reserved
3.12
cleavage
breaking of a mineral along one of its crystallographic directions
3.13
cleavage fragment
fragment of a crystal that is bounded by cleavage (3.12) faces
Note 1 to entry: Crushing of non-asbestiform amphibole generally yields elongated fragments that conform to
the definition of a fibre.
3.14
cluster
structure in which two or more fibres (3.22), or fibre bundles (3.23), are randomly oriented in a
connected grouping
3.15
d-spacing
distance between identical adjacent and parallel planes of atoms in a crystal
3.16
electron diffraction
ED
technique in electron microscopy by which the crystal structure of a specimen is examined
3.17
electron scattering power
extent to which a thin layer of substance scatters electrons from their original directions
3.18
energy dispersive X-ray analysis
EDXA
measurement of the energies and intensities of X-rays by use of a solid-state detector and multi-channel
analyser system
3.19
eucentric
condition when the area of interest of an object is placed on a tilting axis at the intersection of the
electron beam with that axis and is in the plane of focus
3.20
field blank
filter cassette that has been taken to the sampling site, opened and then closed
Note 1 to entry: Such a filter is used to determine the background structure count for the measurement.
3.21
fibril
single fibre (3.22) of asbestos (3.6), which cannot be further separated longitudinally into smaller
components without losing its fibrous properties or appearances
3.22
fibre
elongated particle that has parallel or stepped sides
Note 1 to entry: For the purposes of this document, a fibre is defined to have an aspect ratio equal to or greater
than 5:1 and a minimum length of 0,5 μm.
3.23
fibre bundle
structure composed of parallel, smaller diameter fibres (3.22) attached along their lengths
Note 1 to entry: A fibre bundle may exhibit diverging fibres at one or both ends.
3.24
fibrous structure
fibre, or connected grouping of fibres (3.22), with or without other particles
3.25
habit
characteristic crystal growth form or combination of these forms of a mineral, including characteristic
irregularities
3.26
limit of detection
calculated airborne fibre (3.22) concentration in structures/L, equivalent to the upper 95 % confidence
limit of 2,99 structures predicted by the Poisson distribution for a count of zero structures
3.27
matrix
structure in which one or more fibres (3.22) or fibre bundles (3.23), touch, are attached to, or are
partially concealed by a single particle or connected group of non-fibrous particles
3.28
Miller index
set of either three or four integer numbers used to specify the orientation of a crystallographic plane in
relation to the crystal axes
3.29
phase contrast optical microscopy equivalent fibre
PCM equivalent fibre
fibre (3.22) of aspect ratio greater than or equal to 3:1, longer than 5 μm, and which has a diameter
between 0,2 μm and 3,0 μm
3.30
phase contrast optical microscopy equivalent structure
PCM equivalent structure
fibrous structure (3.24) of aspect ratio greater than or equal to 3:1, longer than 5 μm, and which has a
diameter between 0,2 μm and 3,0 μm
3.31
pixel
smallest image-forming element to which a grey level is assigned
[SOURCE: ISO 23900-6:2015, 2.10]
3.32
primary structure
fibrous structure (3.24) that is a separate entity in the transmission electron microscope image
3.33
replication
procedure in electron microscopy specimen preparation in which a thin copy, or replica, of a surface is
made
3.34
selected area electron diffraction
SAED
technique in electron microscopy in which the crystal structure of a small area of a sample is examined
4 © ISO 2019 – All rights reserved
3.35
serpentine
group of common rock-forming minerals having the nominal formula:
Mg Si O (OH)
3 2 5 4
3.36
structure
single fibre (3.22), fibre bundle (3.23), cluster (3.14) or matrix (3.27)
3.37
twinning
occurrence of crystals of the same species joined together at a particular mutual orientation, and such
that the relative orientations are related by a definite law
3.38
unopened fibre
large diameter asbestos (3.6) fibre bundle (3.23) that has not been separated into its constituent fibrils
or fibres (3.22)
3.39
zone-axis
line or crystallographic direction through the centre of a crystal, which is parallel to the intersection
edges of the crystal faces defining the crystal zone
4 Symbols and abbreviated terms
eV electron volt
kV Kilovolt
l/min litres per minute
−6
μg microgram (10 grams)
−6
μm micrometre (10 metre)
−9
nm nanometre (10 metre)
W Watt
DMF dimethylformamide
ED electron diffraction
EDXA energy dispersive X-ray analysis
FWHM full width, half maximum
HEPA high efficiency particle absolute
MEC mixed esters of cellulose
PC polycarbonate
PCM phase contrast optical microscopy
SAED selected area electron diffraction
SEM scanning electron microscope
STEM scanning transmission electron microscope
TEM transmission electron microscope
UICC Union Internationale Contre le Cancer
5 Type of sample
The method is defined for polycarbonate capillary-pore filters or cellulose ester (either mixed esters of
cellulose or cellulose nitrate) filters through which a known volume of air has been drawn.
6 Range
The upper range of concentration which can be determined is 7 000 structures/mm on the filter.
The lower range is dependent on the area of the TEM specimens analysed, but measurement of
concentrations lower than 1 structure/mm can be achieved. The air concentrations represented by
these values are a function of the volume of air sampled. There is no lower limit to the dimensions of
asbestos fibres which can be detected. In practice, microscopists vary in their ability to detect very
short asbestos fibres. Therefore, a minimum length of 0,5 μm has been defined as the shortest fibre to
be incorporated in the reported results.
The method also includes provision for measurement of the concentrations of fibres with sizes thought
to be of particular biological importance (fibres and bundles >5 µm), and also fibres of sizes defined in
regulations (PCM equivalent fibres).
7 Limit of detection
The limit of detection theoretically can be lowered indefinitely by filtration of progressively larger
volumes of air and by extending the examination of the specimens in the electron microscope. In
practice, the lowest achievable limit of detection for a particular area of TEM specimen examined is
controlled by the total suspended particulate concentration.
For total suspended particulate concentrations of approximately 10 μg/m , corresponding to clean,
rural atmospheres, and assuming filtration of 4 000 l of air, an analytical sensitivity of 0,5 structure/I
can be obtained, equivalent to a limit of detection of 1,8 structure/I, if an area of 0,195 mm of the
TEM specimens is examined. For fibres longer than 5 µm, examined at lower magnifications, this limit
of detection can be reduced by a further order of magnitude. If higher total suspended particulate
concentrations are present, the volume of air filtered must be reduced in order to maintain an acceptable
particulate loading on the filter, leading to a proportionate increase in the analytical sensitivity.
Where this is the case, lower limits of detection can be achieved by increasing the area of the TEM
specimens that is examined. In order to achieve lower limits of detection for fibres and bundles longer
than 5 μm, and for PCM equivalent fibres, lower magnifications are specified which permit more
rapid examination of larger areas of the TEM specimens when the examination is limited to these
dimensions of fibre. The direct analytical method becomes increasingly difficult and imprecise as the
general particulate loading of the sample collection filter increases. It is recommended that no more
than approximately 25 % of the area of the grid openings be occupied by particulate that is capable
of obscuring fibres of interest, which corresponds to approximately 25 µg/cm of filter surface. The
dimensions of the airborne particles on the filter and the dimensional range of fibres being evaluated
determine the extent to which any asbestos could be overlain and obscured. If the total suspended
particulate is largely organic material, the limit of detection can be lowered significantly by using an
indirect preparation method.
6 © ISO 2019 – All rights reserved
8 Principle
A sample of airborne particulate is collected by drawing a measured volume of air through either a
capillary-pore polycarbonate membrane filter of maximum pore size 0,4 µm or a cellulose ester (either
mixed esters of cellulose or cellulose nitrate) membrane filter of maximum pore size 0,45 µm by means
of a battery-powered or mains-powered pump. TEM specimens are prepared from polycarbonate filters
[11]
by a carbon replication procedure in which a thin film of carbon is applied to the filter surface by
vacuum evaporation. Small areas are cut from the carbon-coated filter, supported on TEM specimen
grids, and the filter medium is dissolved away by a solvent extraction procedure. This procedure leaves
a thin film carbon replica of the filter surface, which bridges the openings in the TEM specimen grid, and
which supports each particle from the original filter in its original position. Cellulose ester filters are
chemically treated to collapse the pore structure of the filter, and the surface of the collapsed filter is
[23] [12]
then etched in an oxygen plasma to ensure that all particles are exposed . A thin film of carbon is
evaporated onto the filter surface and small areas are cut from the filter. These sections are supported
[26]
on TEM specimen grids and the filter medium is dissolved away by a solvent extraction procedure .
The TEM specimen grids from either preparation method are examined at both low and high
magnifications to check that they are suitable for analysis before carrying out a quantitative structure
[7]
count on randomly selected grid openings. In the TEM analysis, electron diffraction (ED) is used
to examine the crystal structure of a fibre, and its elemental composition is determined by energy
[6]
dispersive X-ray analysis (EDXA) . For a number of reasons, it is not possible to identify each fibre
unequivocally, and fibres are classified according to the techniques which have been used to identify
them. A simple code is used to record, for each fibre, the manner in which it was classified. The fibre
classification procedure is based on successive inspection of the morphology, the electron diffraction
pattern for a selected area, and the qualitative and quantitative energy dispersive X-ray analyses.
Confirmation of the identification of chrysotile is done only by quantitative ED, and confirmation of
amphibole is done only by quantitative EDXA and quantitative zone axis ED.
In addition to isolated fibres, ambient air samples often contain more complex aggregates of fibres,
with or without other particles. Some particles are composites of asbestos fibres with other materials.
Individual fibres and structures that are more complex are referred to as “asbestos structures”. A
coding system is used to record the type of fibrous structure, and to provide the optimum description
of each of these complex structures. The two codes remove the requirement to interpret the structure
counting data from the microscopist and allow this evaluation to be made later without the requirement
for re-examination of the TEM specimens. Several levels of analysis are specified, the higher levels
providing a more rigorous approach to the identification of fibres. The procedure permits a minimum
required fibre identification criterion to be defined on the basis of previous knowledge, or lack of it,
about the particular sample. Attempts are then made to achieve this minimum criterion for each fibre,
and the degree of success is recorded for each fibre. The lengths and widths of all classified structures
and fibres are recorded. The number of asbestos structures found on a known area of the microscope
sample, together with the equivalent volume of air filtered through this area, is used to calculate the
airborne concentration in asbestos structures/litre of air.
9 Reagents
During the analysis, unless otherwise stated, use only reagents of recognized analytical grade and
water (9.1).
9.1 Water, fibre-free.
A supply of freshly distilled, fibre-free water, or another source of fibre-free, pyrogen-free water shall be
used. Freshly-distilled water, filtered through a 0,1 μm pore size MEC filter, has been found satisfactory.
9.2 Chloroform.
Analytical grade, distilled in glass, preserved with a volume fraction of 1 % ethanol.
9.3 1-Methyl-2-pyrrolidone.
9.4 1,2-Diaminoethane (Ethylene diamine).
9.5 Dimethylformamide.
9.6 Glacial acetic acid.
9.7 Acetone.
10 Apparatus
10.1 Air sampling — Equipment and consumable supplies
10.1.1 Filter cassette
Field monitors, comprising 25 mm to 50 mm diameter conductive three-piece cassettes shall be used
for sample collection. The cassette shall be loaded with either a capillary pore polycarbonate filter of
maximum pore size 0,4 μm or an MEC or cellulose nitrate filter of maximum pore size 0,45 μm. If only
fibres longer than 5 µm are to be included in the measurement, PC filters or MEC filters of maximum
pore size 0,8 µm are permitted. Either type of filter shall be backed by a 5 μm pore size MEC or cellulose
nitrate filter, and supported by a cellulose pad. If push-fit cassettes are used, when the filters are in
position, an elastic cellulose band or adhesive tape shall be applied to prevent air leakage. Suitable
precautions shall be taken to ensure that the filters are tightly clamped in the assembly, so that
significant air leakage around the filter cannot occur.
Representative filters from the filter lot shall be analysed as specified in 12.7 for the presence of
asbestos structures before any are used for air sample collection.
10.1.2 Sampling pump
The sampling pump shall be capable of a flow-rate sufficient to achieve the desired analytical sensitivity.
The face velocity through the filter shall be between 4,0 cm/s and 87,0 cm/s. The sampling pump used
shall provide a non-fluctuating airflow through the filter, and shall maintain the initial volume flow-rate
to within ±10 % throughout the sampling period. A constant flow or critical orifice-controlled pump
meets these requirements. Flexible tubing shall be used to connect the filter cassette to the sampling
pump. A means for calibration of the flow-rate of each pump is also required.
NOTE Some combinations of filter pore size and face velocity can result in distortion of the filter by the
differential pressure across the filter.
10.1.3 Stand
For static sampling, a stand shall be used to hold the filter cassette at the desired height for sampling,
and shall be isolated from the vibrations of the pump (10.1.2).
10.1.4 Personal sampling
For collection of air samples intended to represent the exposure of an individual, the filter cassette
shall be attached within the breathing zone of the individual, i.e. within 25 cm of the worker’s nose
and mouth. Air sampling filter cassettes may be attached to a collar or lapel, with the open end of the
cassette facing down
...
INTERNATIONAL ISO
STANDARD 10312
Second edition
2019-10
Ambient air — Determination of
asbestos fibres — Direct transfer
transmission electron microscopy
method
Air ambiant — Dosage des fibres d'amiante — Méthode par
microscopie électronique à transmission par transfert direct
Reference number
©
ISO 2019
© ISO 2019
All rights reserved. Unless otherwise specified, or required in the context of its implementation, no part of this publication may
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Published in Switzerland
ii © ISO 2019 – All rights reserved
Contents Page
Foreword .v
Introduction .vi
1 Scope . 1
2 Normative references . 1
3 Terms and definitions . 1
4 Symbols and abbreviated terms . 5
5 Type of sample . 6
6 Range . 6
7 Limit of detection . 6
8 Principle . 7
9 Reagents . 7
10 Apparatus . 8
10.1 Air sampling — Equipment and consumable supplies . 8
10.1.1 Filter cassette . 8
10.1.2 Sampling pump . 8
10.1.3 Stand . 8
10.1.4 Personal sampling . 8
10.1.5 Flowmeter . 8
10.2 Specimen preparation laboratory . 9
10.3 Equipment for analysis . 9
10.3.1 Transmission electron microscope . 9
10.3.2 Energy dispersive X-ray analyser .11
10.3.3 Plasma asher .11
10.3.4 Vacuum coating unit .11
10.3.5 Sputter coater .11
10.3.6 Solvent washer (Jaffe washer) .11
10.3.7 Condensation washer .12
10.3.8 Slide warmer or oven .13
10.3.9 Ultrasonic bath .13
10.3.10 Carbon grating replica.13
10.3.11 Calibration specimen grids for EDXA .13
10.3.12 Carbon rod sharpener .14
10.3.13 Disposable tip micropipettes .14
10.4 Consumable supplies .14
10.4.1 Copper or nickel electron microscope grids .14
10.4.2 Gold or nickel electron microscope grids .14
10.4.3 Carbon rod electrodes .14
10.4.4 Routine electron microscopy tools and supplies .14
10.4.5 Reference asbestos samples . .14
10.4.6 Reference samples of mineral fibres other than asbestos .15
11 Air sample collection .15
12 Procedure for analysis .16
12.1 General .16
12.2 Cleaning of sample cassettes .16
12.3 Direct preparation of TEM specimens from polycarbonate filters .17
12.3.1 Selection of filter area for carbon coating .17
12.3.2 Carbon coating of filter portions .17
12.3.3 Preparation of the Jaffe washer .17
12.3.4 Placing of specimens in the Jaffe washer .17
12.4 Direct preparation of TEM specimens from cellulose ester filters .18
12.4.1 Selection of area of filter for preparation .18
12.4.2 Preparation of solution for collapsing cellulose ester filters .18
12.4.3 Filter collapsing procedure .18
12.4.4 Plasma etching of the filter surfaces .18
12.4.5 Carbon coating .18
12.4.6 Preparation of the Jaffe washer .18
12.4.7 Placing of specimens in the Jaffe washer .18
12.5 Criteria for acceptable TEM specimen grids .19
12.6 Procedure for structure counting by TEM.19
12.6.1 General.19
12.6.2 Measurement of mean opening area .20
12.6.3 TEM alignment and calibration procedures .20
12.6.4 Determination of criterion for termination of TEM examination.20
12.6.5 General procedure for structure counting and size analysis .21
12.6.6 Magnification requirements .23
12.7 Blank and quality control determinations .24
12.8 Calculation of results .24
13 Performance characteristics .25
13.1 General .25
13.2 Interferences and limitations of fibre identification .25
13.3 Precision and accuracy.25
13.3.1 Precision.25
13.3.2 Accuracy .26
13.3.3 Inter-laboratory and intra-laboratory analyses .26
13.4 Limit of detection .26
14 Test report .27
Annex A (normative) Determination of operating conditions for plasma asher .30
Annex B (normative) Calibration procedures.31
Annex C (normative) Structure counting criteria .34
Annex D (normative) Fibre identification procedure .44
Annex E (normative) Determination of the concentration of asbestos fibres and bundles
longer than 5 µm, and PCM equivalent asbestos fibres .61
Annex F (normative) Calculation of results .62
Annex G (informative) Strategies for collection of air samples .68
Annex H (informative) Methods for removal of gypsum fibres .69
Bibliography .70
iv © ISO 2019 – All rights reserved
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 146, Air quality, Subcommittee SC 3,
Ambient atmospheres.
This second edition cancels and replaces the first edition (ISO 10312:1995), which has been technically
revised. The main changes compared to the previous edition are as follows:
— the use of electronic display systems with measurement software is permitted;
— the maximum particulate loading for TEM specimens is increased from 10 % to 25 %;
— a simplified fibre identification procedure for investigation of known sources of the regulated
asbestos varieties and richterite/winchite asbestos is permitted;
— the reporting requirements have been changed to permit reporting of the concentrations of fibres
and bundles longer than 5 µm and/or the concentrations of PCM equivalent fibres without the
requirement to report the concentrations of structures equal to or greater than 0,5 µm;
— there is no requirement to report the 95% confidence intervals of the fibre concentrations.
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.
Introduction
This document is applicable to the determination of airborne asbestos in a wide range of ambient
air situations, including the interior atmospheres of buildings, and for a detailed evaluation of
any atmosphere. Because the best available medical evidence indicates that the numerical fibre
concentration and the fibre sizes are the relevant parameters for evaluation of the inhalation hazards,
a fibre counting technique is the only logical approach. Most fibres in ambient atmospheres are not
asbestos and therefore, there is a requirement for fibres to be identified. Many airborne asbestos
fibres in ambient atmospheres have diameters below the resolution limit of the optical microscope.
This document is based on transmission electron microscopy, which has adequate resolution to allow
detection of small fibres and is currently the only technique capable of unequivocal identification of
the majority of individual fibres of asbestos. Airborne asbestos is often found as a mixture of single
fibres and more complex, aggregated structures which may or may not be also aggregated with other
particles. The fibres found suspended in an ambient atmosphere can often be identified unequivocally,
if a sufficient measurement effort is expended. However, if each fibre were to be identified in this way,
the analysis would become prohibitively expensive. Because of instrumental deficiencies or because
of the nature of the particulate, some fibres cannot be positively identified as asbestos, even though
the measurements all indicate that they could be asbestos. Subjective factors therefore contribute
to this measurement, and consequently a very precise definition of the procedure for identification
and enumeration of, asbestos fibres is required. The method specified in this document is designed
to provide the best description possible of the nature, numerical concentration, and sizes of asbestos-
containing particles found in an air sample. This document requires that a very detailed and logical
procedure be used to reduce the subjective aspects of the measurement. The method of data recording
specified in this document is designed to allow re-evaluation of the structure counting data as new
medical evidence becomes available. All feasible specimen preparation techniques result in some
modification of the airborne particulate. Even the collection of particles from a three-dimensional
airborne dispersion onto a two-dimensional filter surface can be considered a modification of the
particulate, and some of the particles in most samples are modified by the specimen preparation
procedures. However, the procedures specified in this document are designed to minimize the
disturbance of the collected particulate material, and the effect of those disturbances that do occur can
be evaluated.
This document describes the method of analysis for a single air filter. However, one of the largest
potential errors in characterizing asbestos in ambient atmospheres is associated with the variability
between filter samples. For this reason, it is necessary to design a replicate sampling scheme in order to
determine this document’s accuracy and precision.
vi © ISO 2019 – All rights reserved
INTERNATIONAL STANDARD ISO 10312:2019(E)
Ambient air — Determination of asbestos fibres — Direct
transfer transmission electron microscopy method
1 Scope
This document specifies a reference method using transmission electron microscopy for the
determination of airborne asbestos fibres and structures in in a wide range of ambient air situations,
including the interior atmospheres of buildings, and for a detailed evaluation for asbestos structures
in any atmosphere. The method allows determination of the type(s) of asbestos fibres present and also
includes measurement of the lengths, widths and aspect ratios of the asbestos structures. The method
cannot discriminate between individual fibres of asbestos and elongate fragments (cleavage fragments
[13]
and acicular particles) from non-asbestos analogues of the same amphibole mineral .
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 4225, Air quality — General aspects — Vocabulary
3 Terms and definitions
For the purposes of this document, the terms and definitions given in ISO 4225 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
acicular
shape shown by an extremely slender crystal with cross-sectional dimensions, which are small relative
to its length, i.e. needle-like
3.2
amphibole
group of rock-forming ferromagnesium silicate minerals, closely related in crystal form and
composition, and having the nominal formula:
A B C T O (OH,F,Cl)
0-1 2 5 8 22 2
where
A = K, Na;
2+
B = Fe , Mn, Mg, Ca, Na;
3+ 2+
C = Al, Cr, Ti, Fe , Mg, Fe ;
3+
T = Si, Al, Cr, Fe , Ti.
Note 1 to entry: In some varieties of amphibole, these elements can be partially substituted by Li, Pb, or Zn.
Amphibole is characterized by a cross-linked double chain of Si-O tetrahedra with a silicon: oxygen ratio of 4:11,
by columnar or fibrous prismatic crystals and by good prismatic cleavage in two directions parallel to the crystal
faces and intersecting at angles of about 56° and 124°.
3.3
amphibole asbestos
amphibole (3.2) in an asbestiform (3.5) habit
3.4
analytical sensitivity
calculated airborne asbestos structure (3.7) concentration in structures/litre, equivalent to counting of
one asbestos (3.6) structure in the analysis
Note 1 to entry: It is expressed in structures/litre.
Note 2 to entry: This method does not specify a unique analytical sensitivity. The analytical sensitivity is
determined by the needs of the measurement and the conditions found on the prepared sample.
3.5
asbestiform
specific type of mineral fibrosity in which the fibres (3.22) and fibrils possess high tensile strength and
flexibility
3.6
asbestos
group of silicate minerals belonging to the serpentine and amphibole (3.2) groups, which have
crystallized in the asbestiform habit, causing them to be easily separated into long, thin, flexible, strong
fibres (3.22) when crushed or processed
Note 1 to entry: The Chemical Abstracts Service Registry Numbers of the most common asbestos varieties are:
chrysotile (12001-29-5), crocidolite (12001-28-4), grunerite asbestos (Amosite) (12172-73-5), anthophyllite
asbestos (77536-67-5), tremolite asbestos (77536-68-6) and actinolite asbestos (77536-66-4). Other varieties of
[19]
asbestiform amphibole, such as richterite asbestos and winchite asbestos may also be found in some products
such as vermiculite and talc.
3.7
asbestos structure
individual fibre (3.22), or any connected or overlapping grouping of asbestos (3.6) fibres or bundles,
with or without other particles
3.8
aspect ratio
ratio of length to width of a particle
3.9
blank
structure count made on transmission electron microscope specimens prepared from an unused filter,
to determine the background measurement
3.10
camera length
equivalent projection length between the specimen and its electron diffraction pattern, in the absence
of lens action
3.11
chrysotile
fibrous mineral of the serpentine group, which has the nominal composition: Mg Si O (OH)
3 2 5 4
Note 1 to entry: Most natural chrysotile deviates little from this nominal composition. In some varieties of
3+ 3+ 2+ 3+
chrysotile, minor substitution of silicon by Al may occur. Minor substitution of magnesium by Al , Fe , Fe ,
2+ 2+ 2+
Ni , Mn and Co may also be present. Chrysotile is the most prevalent type of asbestos.
2 © ISO 2019 – All rights reserved
3.12
cleavage
breaking of a mineral along one of its crystallographic directions
3.13
cleavage fragment
fragment of a crystal that is bounded by cleavage (3.12) faces
Note 1 to entry: Crushing of non-asbestiform amphibole generally yields elongated fragments that conform to
the definition of a fibre.
3.14
cluster
structure in which two or more fibres (3.22), or fibre bundles (3.23), are randomly oriented in a
connected grouping
3.15
d-spacing
distance between identical adjacent and parallel planes of atoms in a crystal
3.16
electron diffraction
ED
technique in electron microscopy by which the crystal structure of a specimen is examined
3.17
electron scattering power
extent to which a thin layer of substance scatters electrons from their original directions
3.18
energy dispersive X-ray analysis
EDXA
measurement of the energies and intensities of X-rays by use of a solid-state detector and multi-channel
analyser system
3.19
eucentric
condition when the area of interest of an object is placed on a tilting axis at the intersection of the
electron beam with that axis and is in the plane of focus
3.20
field blank
filter cassette that has been taken to the sampling site, opened and then closed
Note 1 to entry: Such a filter is used to determine the background structure count for the measurement.
3.21
fibril
single fibre (3.22) of asbestos (3.6), which cannot be further separated longitudinally into smaller
components without losing its fibrous properties or appearances
3.22
fibre
elongated particle that has parallel or stepped sides
Note 1 to entry: For the purposes of this document, a fibre is defined to have an aspect ratio equal to or greater
than 5:1 and a minimum length of 0,5 μm.
3.23
fibre bundle
structure composed of parallel, smaller diameter fibres (3.22) attached along their lengths
Note 1 to entry: A fibre bundle may exhibit diverging fibres at one or both ends.
3.24
fibrous structure
fibre, or connected grouping of fibres (3.22), with or without other particles
3.25
habit
characteristic crystal growth form or combination of these forms of a mineral, including characteristic
irregularities
3.26
limit of detection
calculated airborne fibre (3.22) concentration in structures/L, equivalent to the upper 95 % confidence
limit of 2,99 structures predicted by the Poisson distribution for a count of zero structures
3.27
matrix
structure in which one or more fibres (3.22) or fibre bundles (3.23), touch, are attached to, or are
partially concealed by a single particle or connected group of non-fibrous particles
3.28
Miller index
set of either three or four integer numbers used to specify the orientation of a crystallographic plane in
relation to the crystal axes
3.29
phase contrast optical microscopy equivalent fibre
PCM equivalent fibre
fibre (3.22) of aspect ratio greater than or equal to 3:1, longer than 5 μm, and which has a diameter
between 0,2 μm and 3,0 μm
3.30
phase contrast optical microscopy equivalent structure
PCM equivalent structure
fibrous structure (3.24) of aspect ratio greater than or equal to 3:1, longer than 5 μm, and which has a
diameter between 0,2 μm and 3,0 μm
3.31
pixel
smallest image-forming element to which a grey level is assigned
[SOURCE: ISO 23900-6:2015, 2.10]
3.32
primary structure
fibrous structure (3.24) that is a separate entity in the transmission electron microscope image
3.33
replication
procedure in electron microscopy specimen preparation in which a thin copy, or replica, of a surface is
made
3.34
selected area electron diffraction
SAED
technique in electron microscopy in which the crystal structure of a small area of a sample is examined
4 © ISO 2019 – All rights reserved
3.35
serpentine
group of common rock-forming minerals having the nominal formula:
Mg Si O (OH)
3 2 5 4
3.36
structure
single fibre (3.22), fibre bundle (3.23), cluster (3.14) or matrix (3.27)
3.37
twinning
occurrence of crystals of the same species joined together at a particular mutual orientation, and such
that the relative orientations are related by a definite law
3.38
unopened fibre
large diameter asbestos (3.6) fibre bundle (3.23) that has not been separated into its constituent fibrils
or fibres (3.22)
3.39
zone-axis
line or crystallographic direction through the centre of a crystal, which is parallel to the intersection
edges of the crystal faces defining the crystal zone
4 Symbols and abbreviated terms
eV electron volt
kV Kilovolt
l/min litres per minute
−6
μg microgram (10 grams)
−6
μm micrometre (10 metre)
−9
nm nanometre (10 metre)
W Watt
DMF dimethylformamide
ED electron diffraction
EDXA energy dispersive X-ray analysis
FWHM full width, half maximum
HEPA high efficiency particle absolute
MEC mixed esters of cellulose
PC polycarbonate
PCM phase contrast optical microscopy
SAED selected area electron diffraction
SEM scanning electron microscope
STEM scanning transmission electron microscope
TEM transmission electron microscope
UICC Union Internationale Contre le Cancer
5 Type of sample
The method is defined for polycarbonate capillary-pore filters or cellulose ester (either mixed esters of
cellulose or cellulose nitrate) filters through which a known volume of air has been drawn.
6 Range
The upper range of concentration which can be determined is 7 000 structures/mm on the filter.
The lower range is dependent on the area of the TEM specimens analysed, but measurement of
concentrations lower than 1 structure/mm can be achieved. The air concentrations represented by
these values are a function of the volume of air sampled. There is no lower limit to the dimensions of
asbestos fibres which can be detected. In practice, microscopists vary in their ability to detect very
short asbestos fibres. Therefore, a minimum length of 0,5 μm has been defined as the shortest fibre to
be incorporated in the reported results.
The method also includes provision for measurement of the concentrations of fibres with sizes thought
to be of particular biological importance (fibres and bundles >5 µm), and also fibres of sizes defined in
regulations (PCM equivalent fibres).
7 Limit of detection
The limit of detection theoretically can be lowered indefinitely by filtration of progressively larger
volumes of air and by extending the examination of the specimens in the electron microscope. In
practice, the lowest achievable limit of detection for a particular area of TEM specimen examined is
controlled by the total suspended particulate concentration.
For total suspended particulate concentrations of approximately 10 μg/m , corresponding to clean,
rural atmospheres, and assuming filtration of 4 000 l of air, an analytical sensitivity of 0,5 structure/I
can be obtained, equivalent to a limit of detection of 1,8 structure/I, if an area of 0,195 mm of the
TEM specimens is examined. For fibres longer than 5 µm, examined at lower magnifications, this limit
of detection can be reduced by a further order of magnitude. If higher total suspended particulate
concentrations are present, the volume of air filtered must be reduced in order to maintain an acceptable
particulate loading on the filter, leading to a proportionate increase in the analytical sensitivity.
Where this is the case, lower limits of detection can be achieved by increasing the area of the TEM
specimens that is examined. In order to achieve lower limits of detection for fibres and bundles longer
than 5 μm, and for PCM equivalent fibres, lower magnifications are specified which permit more
rapid examination of larger areas of the TEM specimens when the examination is limited to these
dimensions of fibre. The direct analytical method becomes increasingly difficult and imprecise as the
general particulate loading of the sample collection filter increases. It is recommended that no more
than approximately 25 % of the area of the grid openings be occupied by particulate that is capable
of obscuring fibres of interest, which corresponds to approximately 25 µg/cm of filter surface. The
dimensions of the airborne particles on the filter and the dimensional range of fibres being evaluated
determine the extent to which any asbestos could be overlain and obscured. If the total suspended
particulate is largely organic material, the limit of detection can be lowered significantly by using an
indirect preparation method.
6 © ISO 2019 – All rights reserved
8 Principle
A sample of airborne particulate is collected by drawing a measured volume of air through either a
capillary-pore polycarbonate membrane filter of maximum pore size 0,4 µm or a cellulose ester (either
mixed esters of cellulose or cellulose nitrate) membrane filter of maximum pore size 0,45 µm by means
of a battery-powered or mains-powered pump. TEM specimens are prepared from polycarbonate filters
[11]
by a carbon replication procedure in which a thin film of carbon is applied to the filter surface by
vacuum evaporation. Small areas are cut from the carbon-coated filter, supported on TEM specimen
grids, and the filter medium is dissolved away by a solvent extraction procedure. This procedure leaves
a thin film carbon replica of the filter surface, which bridges the openings in the TEM specimen grid, and
which supports each particle from the original filter in its original position. Cellulose ester filters are
chemically treated to collapse the pore structure of the filter, and the surface of the collapsed filter is
[23] [12]
then etched in an oxygen plasma to ensure that all particles are exposed . A thin film of carbon is
evaporated onto the filter surface and small areas are cut from the filter. These sections are supported
[26]
on TEM specimen grids and the filter medium is dissolved away by a solvent extraction procedure .
The TEM specimen grids from either preparation method are examined at both low and high
magnifications to check that they are suitable for analysis before carrying out a quantitative structure
[7]
count on randomly selected grid openings. In the TEM analysis, electron diffraction (ED) is used
to examine the crystal structure of a fibre, and its elemental composition is determined by energy
[6]
dispersive X-ray analysis (EDXA) . For a number of reasons, it is not possible to identify each fibre
unequivocally, and fibres are classified according to the techniques which have been used to identify
them. A simple code is used to record, for each fibre, the manner in which it was classified. The fibre
classification procedure is based on successive inspection of the morphology, the electron diffraction
pattern for a selected area, and the qualitative and quantitative energy dispersive X-ray analyses.
Confirmation of the identification of chrysotile is done only by quantitative ED, and confirmation of
amphibole is done only by quantitative EDXA and quantitative zone axis ED.
In addition to isolated fibres, ambient air samples often contain more complex aggregates of fibres,
with or without other particles. Some particles are composites of asbestos fibres with other materials.
Individual fibres and structures that are more complex are referred to as “asbestos structures”. A
coding system is used to record the type of fibrous structure, and to provide the optimum description
of each of these complex structures. The two codes remove the requirement to interpret the structure
counting data from the microscopist and allow this evaluation to be made later without the requirement
for re-examination of the TEM specimens. Several levels of analysis are specified, the higher levels
providing a more rigorous approach to the identification of fibres. The procedure permits a minimum
required fibre identification criterion to be defined on the basis of previous knowledge, or lack of it,
about the particular sample. Attempts are then made to achieve this minimum criterion for each fibre,
and the degree of success is recorded for each fibre. The lengths and widths of all classified structures
and fibres are recorded. The number of asbestos structures found on a known area of the microscope
sample, together with the equivalent volume of air filtered through this area, is used to calculate the
airborne concentration in asbestos structures/litre of air.
9 Reagents
During the analysis, unless otherwise stated, use only reagents of recognized analytical grade and
water (9.1).
9.1 Water, fibre-free.
A supply of freshly distilled, fibre-free water, or another source of fibre-free, pyrogen-free water shall be
used. Freshly-distilled water, filtered through a 0,1 μm pore size MEC filter, has been found satisfactory.
9.2 Chloroform.
Analytical grade, distilled in glass, preserved with a volume fraction of 1 % ethanol.
9.3 1-Methyl-2-pyrrolidone.
9.4 1,2-Diaminoethane (Ethylene diamine).
9.5 Dimethylformamide.
9.6 Glacial acetic acid.
9.7 Acetone.
10 Apparatus
10.1 Air sampling — Equipment and consumable supplies
10.1.1 Filter cassette
Field monitors, comprising 25 mm to 50 mm diameter conductive three-piece cassettes shall be used
for sample collection. The cassette shall be loaded with either a capillary pore polycarbonate filter of
maximum pore size 0,4 μm or an MEC or cellulose nitrate filter of maximum pore size 0,45 μm. If only
fibres longer than 5 µm are to be included in the measurement, PC filters or MEC filters of maximum
pore size 0,8 µm are permitted. Either type of filter shall be backed by a 5 μm pore size MEC or cellulose
nitrate filter, and supported by a cellulose pad. If push-fit cassettes are used, when the filters are in
position, an elastic cellulose band or adhesive tape shall be applied to prevent air leakage. Suitable
precautions shall be taken to ensure that the filters are tightly clamped in the assembly, so that
significant air leakage around the filter cannot occur.
Representative filters from the filter lot shall be analysed as specified in 12.7 for the presence of
asbestos structures before any are used for air sample collection.
10.1.2 Sampling pump
The sampling pump shall be capable of a flow-rate sufficient to achieve the desired analytical sensitivity.
The face velocity through the filter shall be between 4,0 cm/s and 87,0 cm/s. The sampling pump used
shall provide a non-fluctuating airflow through the filter, and shall maintain the initial volume flow-rate
to within ±10 % throughout the sampling period. A constant flow or critical orifice-controlled pump
meets these requirements. Flexible tubing shall be used to connect the filter cassette to the sampling
pump. A means for calibration of the flow-rate of each pump is also required.
NOTE Some combinations of filter pore size and face velocity can result in distortion of the filter by the
differential pressure across the filter.
10.1.3 Stand
For static sampling, a stand shall be used to hold the filter cassette at the desired height for sampling,
and shall be isolated from the vibrations of the pump (10.1.2).
10.1.4 Personal sampling
For collection of air samples intended to represent the exposure of an individual, the filter cassette
shall be attached within the breathing zone of the individual, i.e. within 25 cm of the worker’s nose
and mouth. Air sampling filter cassettes may be attached to a collar or lapel, with the open end of the
cassette facing downwards.
10.1.5 Flowmeter
A calibrated flowmeter with an appropriate range and accurate to within 2,5 % of the indicated flow
rate is required for calibration of the air sampling system.
8 © ISO 2019 – All rights reserved
Ensure that the flowmeter is clean before use in order to avoid transfer of asbestos contamination from
the flowmeter to the sample being collected.
10.2 Specimen preparation laboratory
Asbestos, particularly chrysotile, may be present at trace levels in some laboratory reagents. Many
building materials also contain significant amounts of asbestos or other mineral fibres which may
interfere with the analysis if they are inadvertently introduced during preparation of specimens.
It is most important to ensure that, during preparation, contamination of TEM specimens by any
extraneous asbestos fibres is minimized. All specimen preparation steps shall therefore be performed
in an environment where contamination of the sample is minimized. The primary requirement of the
sample p
...
NORME ISO
INTERNATIONALE 10312
Deuxième édition
2019-10
Air ambiant — Dosage des fibres
d'amiante — Méthode par microscopie
électronique à transmission par
transfert direct
Ambient air — Determination of asbestos fibres — Direct transfer
transmission electron microscopy method
Numéro de référence
©
ISO 2019
DOCUMENT PROTÉGÉ PAR COPYRIGHT
© ISO 2019
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
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Publié en Suisse
ii © ISO 2019 – Tous droits réservés
Sommaire Page
Avant-propos .v
Introduction .vi
1 Domaine d’application . 1
2 Références normatives . 1
3 Termes et définitions . 1
4 Symboles et abréviations . 5
5 Type d’échantillon . 6
6 Plage de mesure . 6
7 Limite de détection . 6
8 Principe . 7
9 Réactifs . 8
10 Appareillage . 8
10.1 Prélèvement d’air — Équipement et consommables . 8
10.1.1 Cassette porte-filtre. 8
10.1.2 Pompe de prélèvement . 9
10.1.3 Support . 9
10.1.4 Prélèvement sur individu . 9
10.1.5 Débitmètre . 9
10.2 Laboratoire de préparation des échantillons . 9
10.3 Équipement d’analyse .10
10.3.1 Microscope électronique à transmission .10
10.3.2 Analyseur en dispersion d’énergie des rayons X .12
10.3.3 Four à plasma.12
10.3.4 Évaporateur sous vide .12
10.3.5 Pulvérisateur cathodique .12
10.3.6 Laveur à solvant (laveur Jaffe) .13
10.3.7 Dissolveur à condensation .13
10.3.8 Plaque chauffante ou étuve .14
10.3.9 Bain à ultrasons .14
10.3.10 Réplique d’un réseau carbone .14
10.3.11 Grilles MET d’étalonnage pour SDEX .14
10.3.12 Aiguiseur d’électrodes en carbone .15
10.3.13 Micropipettes à embout jetable .15
10.4 Consommables .15
10.4.1 Grilles de microscope électronique en cuivre ou en nickel .15
10.4.2 Grilles de microscope électronique en or ou en nickel .15
10.4.3 Électrodes en carbone .15
10.4.4 Outils et fournitures courants pour microscopie électronique .15
10.4.5 Échantillons d’amiante de référence .15
10.4.6 Échantillons de référence de fibres minérales autres que l’amiante .16
11 Prélèvement des échantillons d’air .16
12 Mode opératoire d’analyse .17
12.1 Généralités .17
12.2 Nettoyage des cassettes de prélèvement .18
12.3 Préparation directe d’échantillons MET à partir de filtres en polycarbonate .18
12.3.1 Sélection de la surface du filtre pour le dépôt de carbone .18
12.3.2 Dépôt de carbone sur les parties de filtre .18
12.3.3 Préparation du laveur Jaffe .18
12.3.4 Mise en place des échantillons dans le laveur Jaffe.18
12.4 Préparation directe d’échantillons MET à partir de filtres en esters de cellulose .19
12.4.1 Sélection de la surface du filtre à préparer .19
12.4.2 Préparation de la solution pour réduire les filtres en esters de cellulose .19
12.4.3 Mode opératoire pour la réduction du filtre .19
12.4.4 Décapage plasma des surfaces du filtre .19
12.4.5 Dépôt de carbone .20
12.4.6 Préparation du laveur Jaffe .20
12.4.7 Mise en place des échantillons dans le laveur Jaffe.20
12.5 Critères d’acceptation des grilles d’échantillons MET .20
12.6 Mode opératoire de comptage des structures par MET .21
12.6.1 Généralités .21
12.6.2 Mesurage de la surface moyenne d’ouverture .21
12.6.3 Modes opératoires d’alignement et d’étalonnage du MET .22
12.6.4 Détermination du critère d’arrêt de l’examen au MET .22
12.6.5 Mode opératoire général de comptage et d’analyse des dimensions des
structures .22
12.6.6 Exigences de grossissement .25
12.7 Déterminations des blancs et du contrôle qualité .26
12.8 Calcul des résultats .26
13 Caractéristiques de performance .27
13.1 Généralités .27
13.2 Interférences et limites à l’identification des fibres .27
13.3 Fidélité et exactitude .27
13.3.1 Fidélité .27
13.3.2 Exactitude .28
13.3.3 Analyses inter- et intralaboratoires .28
13.4 Limite de détection .28
14 Rapport d’essai .29
Annexe A (normative) Détermination des conditions de fonctionnement du four à plasma .33
Annexe B (normative) Modes opératoires d’étalonnage .34
Annexe C (normative) Critères de comptage des structures .37
Annexe D (normative) Mode opératoire d’identification des fibres .47
Annexe E (normative) Détermination de la concentration en fibres et faisceaux d’amiante
d’une longueur supérieure à 5 µm, et de la concentration en fibres d’amiante
équivalent MOCP .64
Annexe F (normative) Calcul des résultats .65
Annexe G (informative) Stratégies de prélèvement d’échantillons d’air .71
Annexe H (informative) Méthodes d’élimination des fibres de gypse .72
Bibliographie .73
iv © ISO 2019 – Tous droits réservés
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 élaboré par le comité technique ISO/TC 146, Qualité de l’air, sous-comité SC 3,
Atmosphères ambiantes.
Cette deuxième édition annule et remplace la première édition (ISO 10312:1995), qui a fait l’objet d’une
révision technique. Les principales modifications par rapport à l’édition précédente sont les suivantes:
— l’utilisation de systèmes de visualisation électroniques équipés d’un logiciel de mesure est autorisée;
— la densité de particules maximale pour les échantillons MET est portée de 10 % à 25 %;
— un mode opératoire simplifié d’identification des fibres pour étudier les sources connues des
variétés d’amiante réglementées et d’amiante richtérite/winchite est autorisé;
— les exigences en matière de rapport d’essai ont été modifiées pour permettre de consigner les
concentrations de fibres et de faisceaux de plus de 5 µm de longueur et/ou les concentrations de fibres
équivalent MOCP sans qu’il soit exigé de consigner les concentrations de structures supérieures ou
égales à 0,5 µm;
— il n’est pas exigé de consigner les intervalles de confiance à 95 % des concentrations de fibres.
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.
Introduction
Le présent document est applicable au dosage de l’amiante en suspension dans l’air ambiant dans un
nombre varié de situations, y compris les atmosphères intérieures des bâtiments, et à une évaluation
détaillée de toute atmosphère. Les recherches médicales les plus avancées indiquant que la concentration
numérique des fibres ainsi que leur taille sont les paramètres les plus pertinents pour évaluer les risques
pour la santé liés à l’inhalation, une technique de comptage des fibres constitue la seule approche
logique. La plupart des fibres en suspension dans les atmosphères ambiantes ne sont pas de l’amiante
et, par conséquent, il est nécessaire de les identifier. De nombreuses fibres d’amiante en suspension
dans l’air dans les atmosphères ambiantes ont des diamètres inférieurs à la limite de résolution du
microscope optique. Le présent document est fondé sur la microscopie électronique à transmission, qui
a une résolution adéquate pour permettre la détection de petites fibres et qui est actuellement la seule
technique capable d’identifier sans équivoque la majorité des fibres individuelles d’amiante. L’amiante
en suspension dans l’air se présente souvent sous forme de mélange de fibres individuelles et de
structures agrégées plus complexes qui peuvent également être ou non agrégées à d’autres particules.
Les fibres trouvées en suspension dans une atmosphère ambiante peuvent souvent être identifiées sans
équivoque, si un soin suffisant est apporté à l’analyse. Cependant, si chaque fibre devait être identifiée de
cette manière, le coût de l’analyse deviendrait prohibitif. En raison des insuffisances des instruments ou
de la nature des particules, certaines fibres ne peuvent pas être formellement identifiées comme étant
de l’amiante, même si les mesures indiquent toutes qu’elles pourraient en être. Des facteurs subjectifs
interviennent dans ces mesurages et, par conséquent, une définition très précise du mode opératoire
d’identification et de comptage des fibres d’amiante est nécessaire. La méthode spécifiée dans le présent
document est destinée à fournir la meilleure description possible de la nature, de la concentration
numérique et des tailles des particules contenant de l’amiante trouvées dans un échantillon d’air. Le
présent document exige l’utilisation d’un mode opératoire très détaillé et logique pour réduire les
aspects subjectifs du mesurage. La méthode d’enregistrement des données spécifiée dans le présent
document est destinée à permettre une réévaluation des données de comptage des structures à mesure
que de nouvelles données médicales sont disponibles. Toutes les techniques possibles de préparation
des échantillons entraînent des modifications des caractéristiques des particules en suspension dans
l’air. Le prélèvement même de particules à partir d’une dispersion tridimensionnelle sur la surface d’un
filtre bidimensionnelle peut être considéré comme apportant des modifications aux caractéristiques
des particules; en outre, pour la plupart des échantillons, certaines caractéristiques sont également
modifiées par les modes opératoires de préparation. Toutefois, les modes opératoires spécifiés dans
le présent document sont destinés à réduire au minimum la perturbation de la matière particulaire
recueillie et l’effet des perturbations qui se produisent peut être évalué.
Le présent document décrit la méthode d’analyse applicable à un seul filtre à air. Cependant, l’une des
plus grandes erreurs qui peut se produire lors de la caractérisation de l’amiante dans les atmosphères
ambiantes est associée à la variabilité entre des échantillons de filtre. Pour cette raison, il est nécessaire
de prévoir un plan d’échantillonnage répété afin de déterminer l’exactitude et la fidélité du présent
document.
vi © ISO 2019 – Tous droits réservés
NORME INTERNATIONALE ISO 10312:2019(F)
Air ambiant — Dosage des fibres d'amiante — Méthode
par microscopie électronique à transmission par
transfert direct
1 Domaine d’application
Le présent document spécifie une méthode de référence utilisant la microscopie électronique à
transmission pour déterminer la concentration en fibres et structures d’amiante en suspension dans l’air
dans diverses atmosphères ambiantes, notamment les atmosphères intérieures de bâtiments, et pour
évaluer en détail les structures d’amiante dans les atmosphères. Cette méthode permet de déterminer
le(s) type(s) de fibres d’amiante présentes et comprend également le mesurage des longueurs, des
largeurs et des rapports longueur/largeur des structures d’amiante. Elle ne peut pas faire la différence
entre des fibres individuelles d’amiante et des fragments allongés (fragments de clivage et particules
[13]
aciculaires) d’analogues non asbestiformes du même minéral amphibole .
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).
ISO 4225, Qualité de l’air — Aspects généraux — Vocabulaire
3 Termes et définitions
Pour les besoins du présent document, les termes et définitions de l’ISO 4225 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 http:// www .electropedia .org/ .
3.1
aciculaire
forme d’un cristal extrêmement mince avec une section petite par rapport à sa longueur, par exemple
en forme d’aiguille
3.2
amphibole
groupe de minéraux formés de silicate de fer ou magnésium, étroitement liés sous forme cristalline,
avec la composition chimique nominale:
A B C T O (OH,F,Cl)
0-1 2 5 8 22 2
où
A = K, Na;
2+
B = Fe , Mn, Mg, Ca, Na;
3+ 2+
C = Al, Cr, Ti, Fe , Mg, Fe ;
3+
T = Si, Al, Cr, Fe , Ti
Note 1 à l'article: Dans certaines variétés d’amphibole, ces éléments peuvent être partiellement substitués par Li,
Pb ou Zn. L’amphibole est caractérisée par une double chaîne réticulée formée de tétraèdres Si-O avec un rapport
silicium/oxygène de 4/11, par des cristaux prismatiques en forme de colonne ou de fibre et par un clivage
prismatique en deux directions parallèles à la surface des cristaux et se croisant à des angles d’environ 56°
et 124°.
3.3
amiante amphibole
amphibole (3.2) ayant un faciès asbestiforme (3.5)
3.4
sensibilité analytique
concentration calculée de structures d’amiante (3.7) en suspension par litre d’air, équivalant à
l’observation d’une structure d’amiante (3.6) dans l’analyse
Note 1 à l'article: Elle est exprimée en nombre de structures/litre.
Note 2 à l'article: La présente méthode ne spécifie pas de sensibilité analytique unique. La sensibilité analytique
est déterminée par les besoins du mesurage et par les conditions observées sur l’échantillon préparé.
3.5
asbestiforme
type spécifique de minéral fibreux dans lequel les fibres (3.22) et les fibrilles possèdent une haute
résistance à la traction et une grande souplesse
3.6
amiante
groupe de minéraux de silicates appartenant aux groupes des amphiboles (3.2) et des serpentines, qui
se sont cristallisés en faciès asbestiforme (3.5), ce qui permet, lorsqu’ils sont traités ou broyés, de les
séparer facilement en fibres (3.22) longues, fines, souples et solides
Note 1 à l'article: Les numéros d’enregistrement du Chemical Abstracts Service pour les variétés d’amiante les
plus courantes sont: chrysotile (12001-29-5), crocidolite (12001-28-4), amiante grünérite (amosite) (12172-73-
5), amiante anthophyllite (77536-67-5), amiante trémolite (77536-68-6) et amiante actinolite (77536-66-4).
[19]
D’autres variétés d’amphibole asbestiforme, notamment l’amiante richtérite et l’amiante winchite peuvent
également être présentes dans certains produits tels que la vermiculite et le talc.
3.7
structure d’amiante
fibre (3.22) individuelle ou tout groupement contigu ou formé par chevauchement de fibres ou de
faisceaux d’ amiante (3.6), avec ou sans particules associées
3.8
rapport longueur/largeur
rapport de la longueur d’une particule à sa largeur
2 © ISO 2019 – Tous droits réservés
3.9
blanc
comptage de structures effectué sur des échantillons pour microscopie électronique à transmission
préparés à partir d’un filtre non utilisé pour déterminer la concentration en bruit de fond
3.10
longueur de caméra
longueur de projection équivalente entre l’échantillon et le diagramme de diffraction électronique, en
l’absence d’action d’une lentille
3.11
chrysotile
minéral fibreux du groupe des serpentines, ayant une composition répondant à la formule chimique
brute: Mg Si O (OH)
3 2 5 4
Note 1 à l'article: La plupart des chrysotiles naturels s’écartent peu de cette composition nominale. Dans certaines
3+
variétés, il peut se produire une substitution mineure de silicium par de l’Al . Une substitution mineure de
3+ 2+ 3+ 2+ 2+ 2+
magnésium par de l’Al , du Fe , du Fe , du Ni , du Mn et du Co peut aussi se présenter. Le chrysotile est le
type d’amiante le plus répandu.
3.12
clivage
fracture d’un minéral dans l’une de ses directions cristallographiques
3.13
fragment de clivage
fragment de cristal délimité par les plans de clivage (3.12)
Note 1 à l'article: En général, le broyage de l’amphibole non asbestiforme produit des fragments allongés
conformes à la définition d’une fibre.
3.14
agglomérat
structure dans laquelle deux ou plusieurs fibres (3.22), ou faisceaux de fibres (3.23) sont orientés au
hasard et forment un groupement contigu
3.15
espace interréticulaire
distance entre des plans identiques parallèles et adjacents d’atomes du cristal
3.16
diffraction électronique
ED
technique utilisée en microscopie électronique permettant d’examiner la structure cristalline d’un
échantillon
3.17
pouvoir de diffusion d’électrons
capacité d’une couche mince de substance diffuse à dévier des électrons de leur direction d’origine
3.18
analyse en dispersion d’énergie des rayons X
SDEX
mesurage des énergies et des intensités des rayons X à l’aide d’un détecteur à semi-conducteurs et d’un
système analyseur à voies multiples
3.19
eucentrique
condition d’un objet dont la zone d’observation est placée sur un axe d’inclinaison au point d’intersection
avec le faisceau d’électrons et est dans le plan de focalisation
3.20
témoin
filtre qui a été emporté sur le site de prélèvement, et dont la cassette a été ouverte et refermée
Note 1 à l'article: Un tel filtre sert à déterminer le nombre de structures en bruit de fond.
3.21
fibrille
fibre (3.22) unitaire d’ amiante (3.6) qui ne peut pas être séparée davantage longitudinalement en
composants plus petits sans perdre ses propriétés de fibres ou son apparence
3.22
fibre
particule allongée qui a des côtés parallèles ou étagés
Note 1 à l'article: Aux fins du présent document, une fibre est définie comme ayant un rapport longueur/largeur
supérieur ou égal à 5/1 et une longueur minimale de 0,5 µm.
3.23
faisceau de fibres
structure composée de fibres (3.22) parallèles de diamètres inférieurs attachées sur leur longueur
Note 1 à l'article: Un faisceau de fibres peut présenter des fibres divergentes à l’une ou aux deux extrémités.
3.24
structure fibreuse
fibre ou groupement contigu de fibres (3.22) avec ou sans particules associées
3.25
faciès
forme cristalline caractéristique ou combinaison de ces formes d’un minéral, y compris les irrégularités
caractéristiques
3.26
limite de détection
concentration de fibres (3.22) en suspension dans l’air calculée en structures par litre, équivalant à la
limite supérieure de l’intervalle de confiance à 95 % de 2,99 structures prévue par la loi de Poisson
pour un comptage de zéro structure
3.27
matrice
structure dans laquelle une ou plusieurs fibres (3.22) ou un ou plusieurs faisceaux de fibres (3.23)
en contact sont attaché(e)s à ou partiellement dissimulé(e)s par une particule unitaire ou un groupe
contigu de particules non fibreuses
3.28
indice de Miller
ensemble de trois ou quatre nombres entiers utilisés pour spécifier l’orientation d’un plan
cristallographique par rapport aux axes d’un cristal
3.29
fibre équivalent MOCP
fibre équivalente en microscopie optique en contraste de phase
fibre (3.22) de rapport longueur/largeur supérieur ou égal à 3/1, de longueur supérieure à 5 µm et dont
le diamètre est compris entre 0,2 µm et 3,0 µm
3.30
structure équivalent MOCP
structure équivalente en microscopie optique en contraste de phase
structure fibreuse (3.24) de rapport longueur/largeur supérieur ou égal à 3/1, de longueur supérieure à
5 µm et dont le diamètre est compris entre 0,2 µm et 3,0 µm
4 © ISO 2019 – Tous droits réservés
3.31
pixel
plus petit élément formant une image auquel est assigné un niveau de gris
[SOURCE: ISO 23900-6:2015, 2.10]
3.32
structure primaire
structure fibreuse (3.24) qui représente une entité distincte sur l’image du microscope électronique à
transmission
3.33
réplication
méthode de préparation d’échantillons de microscopie électronique dans laquelle une copie mince ou
réplique d’une surface est faite
3.34
microdiffraction électronique en aire sélectionnée
SAED
technique utilisée en microscopie électronique dans laquelle la structure cristalline d’une petite surface
d’un échantillon est examinée
3.35
serpentine
groupe de minéraux communs de formule chimique brute:
Mg Si O (OH)
3 2 5 4
3.36
structure
fibre (3.22) individuelle, faisceau de fibres (3.23), agglomérat (3.14) ou matrice (3.27)
3.37
maclage
phénomène par lequel des cristaux de même espèce sont accolés ensemble suivant une orientation
particulière de sorte que les orientations relatives sont reliées par une loi bien définie
3.38
fibre non ouverte
faisceau de fibres (3.23) d’ amiante (3.6) de grand diamètre qui n’a pas été divisé en fibrilles ou fibres
(3.22) le constituant
3.39
axe de zone
ligne ou direction cristallographique à travers le centre d’un cristal qui est parallèle aux arêtes
d’intersection des faces d’un cristal définissant la zone cristalline
4 Symboles et abréviations
eV électronvolt
kV kilovolt
l/min litres par minute
−6
μg microgramme (10 gramme)
−6
μm micromètre (10 mètre)
−9
nm nanomètre (10 mètre)
W watt
DMF diméthylformamide
ED diffraction électronique
SDEX analyse en dispersion d’énergie des rayons X
FWHM largeur totale à mi-hauteur
HEPA filtre de haute efficacité pour l’arrêt des particules
MEC esters mélangés de cellulose
PC polycarbonate
MOCP microscopie optique en contraste de phase
SAED microdiffraction électronique en aire sélectionnée
MEB microscope électronique à balayage
MEBT microscope électronique à transmission avec balayage
MET microscope électronique à transmission
UICC Union internationale contre le cancer
5 Type d’échantillon
La méthode est définie pour des filtres en polycarbonate à pores capillaires ou en esters de cellulose
(esters mélangés de cellulose ou nitrate de cellulose) à travers lesquels un volume connu d’air a été aspiré.
6 Plage de mesure
La limite supérieure de la gamme de concentrations qui peut être déterminée sur le filtre est
de 7 000 structures/mm . La limite inférieure de la gamme dépend de la surface des échantillons MET
analysés, mais il est possible de mesurer des concentrations inférieures à 1 structure/mm . Les
concentrations dans l’air représentées par ces valeurs sont fonction du volume d’air prélevé. Il n’existe
pas de limite inférieure applicable aux dimensions des fibres d’amiante qui peuvent être détectées.
Dans la pratique, les techniciens n’ont pas tous la même aptitude à détecter les fibres d’amiante très
courtes. Par conséquent, une longueur minimale de 0,5 µm a été définie comme longueur des fibres les
plus courtes à être intégrée aux résultats consignés.
La méthode prévoit également de mesurer les concentrations de fibres dont les tailles sont jugées avoir
une importance biologique particulière (fibres et faisceaux > 5 µm) ainsi que les fibres dont les tailles
sont définies dans les réglementations (fibres équivalent MOCP).
7 Limite de détection
La limite de détection peut en théorie être abaissée indéfiniment par la filtration de volumes de plus en
plus importants d’air et par le prolongement de l’examen des échantillons au microscope électronique.
Dans la pratique, la limite de détection la plus basse qu’il est possible d’atteindre pour une surface
particulière d’échantillon MET examinée est contrôlée par la concentration totale des particules en
suspension.
6 © ISO 2019 – Tous droits réservés
Pour les concentrations totales de particules en suspension d’environ 10 µg/m , correspondant à des
atmosphères rurales propres, et dans l’hypothèse d’une filtration de 4 000 l d’air, il est possible d’obtenir
une sensibilité analytique de 0,5 structure/l, équivalant à une limite de détection de 1,8 structure/l, si
une surface de 0,195 mm d’échantillons MET est examinée. Pour les fibres de plus de 5 µm de longueur
examinées sous un grossissement moins élevé, cette limite de détection peut être réduite d’un ordre de
grandeur supplémentaire. En cas de concentrations totales de particules en suspension plus élevées, il
faut réduire le volume d’air filtré afin de maintenir une densité de particules acceptables sur le filtre, ce
qui entraîne une augmentation proportionnelle de la sensibilité analytique.
Le cas échéant, des limites de détection moins élevées peuvent être atteintes en augmentant la surface
des échantillons MET examinés. Pour atteindre des limites de détection inférieures pour les fibres
et les faisceaux de plus de 5 µm de longueur, et pour les fibres équivalent MOCP, des grossissements
moins élevés sont spécifiés et permettent d’étudier plus rapidement de plus grandes surfaces
d’échantillons MET lorsque l’examen se limite à ces dimensions de fibres. La méthode d’analyse directe
devient de plus en plus complexe et imprécise à mesure que la densité globale des particules du filtre
de prélèvement d’échantillons augmente. Il est recommandé que la surface des ouvertures de grille
occupée par les particules capables de dissimuler les fibres étudiées ne dépasse pas 25 % environ, ce qui
correspond à environ 25 µg/cm de surface de filtre. Les dimensions des particules en suspension dans
l’air sur le filtre et la plage dimensionnelle des fibres évaluées déterminent le degré de recouvrement
et de dissimulation potentiel de l’amiante. Si les particules totales en suspension sont essentiellement
un matériau organique, la limite de détection peut être abaissée significativement en utilisant une
méthode de préparation indirecte.
8 Principe
Un échantillon de particules en suspension dans l’air est recueilli en aspirant un volume mesuré d’air
à travers soit un filtre à membrane en polycarbonate à pores capillaires d’une porosité maximale
de 0,4 µm, soit un filtre à membrane en esters de cellulose (esters mélangés de cellulose ou nitrate
de cellulose) d’une porosité maximale de 0,45 µm au moyen d’une pompe alimentée sur le secteur ou
par batterie. Les échantillons MET sont préparés à partir des filtres en polycarbonate par un mode
[11]
opératoire de réplication sur carbone dans lequel un mince film de carbone est appliqué sur la
surface du filtre par évaporation sous vide. De petites sections sont découpées dans le filtre carboné,
placées sur des grilles d’échantillons MET et le milieu filtrant est dissous par un mode opératoire
d’extraction au solvant. Ce mode opératoire laisse un mince film de carbone, réplique de la surface
du filtre, qui recouvre les ouvertures de la grille d’échantillon MET et maintient chaque particule du
filtre d’origine dans sa position d’origine. Les filtres en esters de cellulose sont traités chimiquement
pour réduire la structure poreuse du filtre. La surface du filtre réduit est alors décapée par plasma
[23] [12]
d’oxygène pour s’assurer que toutes les particules sont exposées. Un mince film de carbone est
déposé par évaporation à la surface du filtre et de petites sections sont découpées dans le filtre. Ces
sections sont placées sur des grilles d’échantillons MET et le milieu filtrant est dissous par un mode
[26]
opératoire d’extraction au solvant .
Les grilles d’échantillons MET provenant de l’une ou l’autre méthode de préparation sont examinées
à faible et à fort grossissements pour s’assurer qu’elles conviennent pour l’analyse avant d’effectuer
une évaluation quantitative des structures sur des ouvertures de grille choisies au hasard. Lors de
[7]
l’analyse MET, la diffraction électronique (ED) est utilisée pour examiner la structure cristalline
d’une fibre, et sa composition élémentaire est déterminée par une analyse en dispersion d’énergie des
[6]
rayons X (SDEX). Pour un certain nombre de raisons, il n’est pas possible d’identifier sans équivoque
chaque fibre, et les fibres sont classées en fonction des techniques qui ont été utilisées pour les
identifier. Un code simple est utilisé pour consigner pour chaque fibre la manière dont elle a été classée.
Le mode opératoire de classification des fibres est fondé sur un examen successif de la morphologie, du
diagramme de microdiffraction électronique en aire sélectionnée, ainsi que des analyses qualitative et
quantitative en dispersion d’énergie des rayons X. La confirmation de l’identification du chrysotile se
fait uniquement par ED quantitative et celle de l’amphibole se fait uniquement en combinant la SDEX
quantitative et l’ED quantitative en axe de zone.
En plus de fibres isolées, les échantillons d’air ambiant contiennent souvent des agrégats plus complexes
de fibres, associés ou non à d’autres particules. Certaines particules sont composées de fibres
d’amiante et d’autres matériaux. Les fibres individuelles et les structures plus complexes sont appelées
«structures d’amiante». Un système de codage est utilisé pour consigner le type de structure fibreuse
et pour donner la meilleure description de chacune de ces structures complexes. Les deux codes évitent
au technicien la nécessité d’interpréter les données de comptage des structures et permettent de
faire cette évaluation ultérieurement sans avoir besoin de réexaminer les échantillons MET. Plusieurs
niveaux d’analyse sont spécifiés, les niveaux supérieurs fournissant une approche plus rigoureuse pour
l’identification des fibres. Le mode opératoire permet de définir un critère minimal pour l’identification
requise des fibres, fondé sur les connaissances préalables, ou leur absence, concernant l’échantillon
particulier. On s’efforce alors de parvenir à ce critère minimal pour chaque fibre et le degré de réussite
est consigné pour chaque fibre. Les longueurs et les largeurs de toutes les structures et fibres classées
sont consignées. Le nombre de structures d’amiante trouvées sur une surface connue de l’échantillon
observée au microscope, ainsi que le volume équivalent d’air filtré à travers cette surface, sont utilisés
pour calculer la concentration en structures d’amiante en suspension par litre d’air.
9 Réactifs
Pendant l’analyse, sauf mention contraire, utiliser uniquement des réactifs de qualité analytique
reconnue et de l’eau (9.1).
9.1 Eau, exempte de fibres.
Une alimentation en eau récemment distillée et exempte de fibres, ou une autre source d’eau exempte
de fibres et d’agents pyrogènes, doit être utilisée. De l’eau récemment distillée, filtrée à travers un filtre
en esters mélangés de cellulose d’une porosité de 0,1 µm, s’est avérée satisfaisante.
9.2 Chloroforme.
Qualité analytique, distillé dans un récipient en verre, conservé avec de l’éthanol à 1 % (fraction
volumique).
9.3 1-méthyl-2-pyrrolidone
9.4 1,2-diaminoéthane (éthylènediamine)
9.5 Diméthylformamide
9.6 Acide acétique glacial
9.7 Acétone
10 Appareillage
10.1 Prélèvement d’air — Équipement et consommables
10.1.1 Cassette porte-filtre
Des dispositifs de contrôle sur le terrain comprenant des cassettes conductrices en trois
...
Frequently Asked Questions
SIST ISO 10312:2019 is a standard published by the Slovenian Institute for Standardization (SIST). Its full title is "Ambient air - Determination of asbestos fibres - Direct transfer transmission electron microscopy method". This standard covers: This document specifies a reference method using transmission electron microscopy for the determination of airborne asbestos fibres and structures in in a wide range of ambient air situations, including the interior atmospheres of buildings, and for a detailed evaluation for asbestos structures in any atmosphere. The method allows determination of the type(s) of asbestos fibres present and also includes measurement of the lengths, widths and aspect ratios of the asbestos structures. The method cannot discriminate between individual fibres of asbestos and elongate fragments (cleavage fragments and acicular particles) from non-asbestos analogues of the same amphibole mineral[13].
This document specifies a reference method using transmission electron microscopy for the determination of airborne asbestos fibres and structures in in a wide range of ambient air situations, including the interior atmospheres of buildings, and for a detailed evaluation for asbestos structures in any atmosphere. The method allows determination of the type(s) of asbestos fibres present and also includes measurement of the lengths, widths and aspect ratios of the asbestos structures. The method cannot discriminate between individual fibres of asbestos and elongate fragments (cleavage fragments and acicular particles) from non-asbestos analogues of the same amphibole mineral[13].
SIST ISO 10312:2019 is classified under the following ICS (International Classification for Standards) categories: 13.040.20 - Ambient atmospheres. The ICS classification helps identify the subject area and facilitates finding related standards.
SIST ISO 10312:2019 has the following relationships with other standards: It is inter standard links to SIST ISO 10312:1996. Understanding these relationships helps ensure you are using the most current and applicable version of the standard.
SIST ISO 10312:2019 is associated with the following European legislation: EU Directives/Regulations: 2005-01-4036, 2005-01-5153. When a standard is cited in the Official Journal of the European Union, products manufactured in conformity with it benefit from a presumption of conformity with the essential requirements of the corresponding EU directive or regulation.
You can purchase SIST ISO 10312:2019 directly from iTeh Standards. The document is available in PDF format and is delivered instantly after payment. Add the standard to your cart and complete the secure checkout process. iTeh Standards is an authorized distributor of SIST standards.
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