ASTM E2228-23a

This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the
Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.
Designation: E2228 − 23a An American National Standard
Standard Guide for
Microscopical Examination of Textile Fibers
This standard is issued under the fixed designation E2228; the number immediately following the designation indicates the year of
original adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. A
superscript epsilon (´) indicates an editorial change since the last revision or reapproval.
1. Scope D276 Test Methods for Identification of Fibers in Textiles
(Withdrawn 2021)
1.1 This standard covers guidelines for microscopical ex-
E620 Practice for Reporting Opinions of Scientific or Tech-
aminations employed in forensic fiber classification,
nical Experts
identification, and comparison. The microscopical examination
E1459 Guide for Physical Evidence Labeling and Related
of fibers includes the use of a variety of light microscopes, such
Documentation
as stereomicroscopes, compound microscopes, and comparison
E1492 Practice for Receiving, Documenting, Storing, and
microscopes, as well as a variety of illumination types, such as
Retrieving Evidence in a Forensic Science Laboratory
bright field, polarized light, fluorescence, and interference. In
E1732 Terminology Relating to Forensic Science
certain instances, the scanning electron microscope can yield
E2917 Practice for Forensic Science Practitioner Training,
additional information. The particular test(s) or techniques
Continuing Education, and Professional Development
employed by each examiner or laboratory will depend upon
Programs
available equipment, examiner training, and the nature and
2.2 AATCC Standards:
extent of the fiber evidence.
AATCC Test Methods 20 Fiber Identification: Qualitative
1.2 The values stated in SI units are to be regarded as
2.3 Other Documents:
standard. No other units of measurement are included in this
ISO 17025 Testing and calibration laboratories
standard.
3. Terminology
1.3 This standard is intended for use by competent forensic
science practitioners with the requisite formal education, 3.1 Definitions—For definitions of terms used in this guide,
refer to Terminology D123 and E1732.
discipline-specific training (see Practice E2917), and demon-
strated proficiency to perform forensic casework.
3.2 Definitions of Terms Specific to This Standard:
1.4 This standard does not purport to address all of the 3.2.1 anisotropic, adj—a characteristic of an object in which
safety concerns, if any, associated with its use. It is the
the refractive index differs depending on the direction of
responsibility of the user of this standard to establish appro-
propagation or vibration of light through the object.
priate safety, health, and environmental practices and deter-
(1)
mine the applicability of regulatory limitations prior to use.
3.2.2 barrier filter, n—a filter used in fluorescence micros-
1.5 This international standard was developed in accor-
copy that absorbs excitation energy that has been reflected by
dance with internationally recognized principles on standard-
the sample, selectively transmitting only wavelengths of light
ization established in the Decision on Principles for the
greater than the cut-off wavelength, or within a specific
Development of International Standards, Guides and Recom-
wavelength range.
mendations issued by the World Trade Organization Technical
3.2.3 Becke line, n—the bright halo near the boundary of a
Barriers to Trade (TBT) Committee.
fiber that moves with respect to that boundary as the micro-
2. Referenced Documents scope is focused through best focus when the fiber is mounted
in a medium that differs from its refractive index.
2.1 ASTM Standards:
(1)
D123 Terminology Relating to Textiles
1 3
This guide is under the jurisdiction of ASTM Committee E30 on Forensic The last approved version of this historical standard is referenced on
Sciences and is the direct responsibility of Subcommittee E30.01 on Criminalistics. www.astm.org.
Current edition approved May 1, 2023. Published May 2023. Originally Available from American Association of Textile Chemists and Colorists
approved in 2002. Last previous edition approved in 2023 as E2228 – 23. DOI: (AATCC), P.O. Box 12215, Research Triangle Park, NC 27709-2215, http://
10.1520/E2228-23A. www.aatcc.org.
2 5
For referenced ASTM standards, visit the ASTM website, www.astm.org, or Available from American National Standards Institute (ANSI), 25 W. 43rd St.,
contact ASTM Customer Service at service@astm.org. For Annual Book of ASTM 4th Floor, New York, NY 10036, http://www.ansi.org.
Standards volume information, refer to the standard’s Document Summary page on The boldface numbers in parentheses refer to a list of references at the end of
the ASTM website. this standard.
Copyright © ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States
E2228 − 23a
3.2.4 Becke line method, n—a method for determining the 3.2.13 cortex, n—the main structural component of hair
refractive index of a fiber relative to its mountant by noting the consisting of elongated and fusiform (spindle-shaped) cells; the
direction in which the Becke line moves when the focus is cortex can contain pigment granules, air spaces called cortical
changed. fusi, and structures called ovoid bodies.
(1)
3.2.14 crimp, n—the curl, wave, or compression that is
3.2.4.1 Discussion—The Becke line always moves toward
naturally occurring or otherwise imparted to a fiber.
the higher refractive index medium (fiber or mountant) when
3.2.15 cuticle, n—in mammalian hair fibers, the layers of
focus is raised (stage is lowered) and towards the lower
flattened cells enclosing the cortex, which form an envelope of
refractive index medium when focus is lowered (stage is
overlapping scales surrounding the fiber. D123
raised). At the point where the index of the fiber matches the
index of the mounting medium, the Becke line is no longer
3.2.16 delustrant, n—a pigment, usually titanium dioxide,
visible. The Becke line is generally viewed at a wavelength of used to dull the luster of a manufactured fiber.
589 nm (the D line of Sodium [n ]).
(3)
D
(1)
3.2.17 dichroism, n—the property of exhibiting different
3.2.5 birefringence, n—the numerical difference in refrac-
colors, especially two different colors, when viewed along
tive indices (n) for a fiber, given by the equation:
different axes by plane polarized light.
ni2n'?
3.2.18 dislocations, n—distinct features that occur in natural
?
fibers (for example, flax, ramie, jute, hemp) in the shape of X’s,
Birefringence (B) can be calculated by determining the
I’s, and V’s that are present along the fiber cell wall; these
retardation (r) and thickness (T) at a particular point in a
features are often useful for identification.
fiber and by using the equation:
3.2.19 dispersion of birefringence, n—the variation of bire-
B 5 r ~nm!⁄1000T ~µm!
fringence with wavelength of light.
(1)
3.2.19.1 Discussion—When dispersion of birefringence is
3.2.6 comparison microscope, n—a system of two micro- significant in a particular fiber, anomalous interference colors
scopes positioned side-by-side and connected via an optical not appearing in the regular color sequence of the Michel-Lévy
bridge so that two specimens are examined simultaneously in chart can result. Strong dispersion of birefringence can also
a single field of view in either transmitted or reflected light. interfere with the accurate determination of retardation in
highly birefringent fibers.
3.2.7 compensator, n—any variety of optical devices that
can be placed in the light path of a polarized light microscope
3.2.20 dispersion staining, n—an optical staining technique
to introduce known, fixed or variable retardation in a specific
in which colors are produced by the differential refraction of
vibration direction; the retardation and sign of elongation of the
different wavelengths of light due to mounting the sample in a
fiber can then be determined.
liquid having a different dispersion of refractive index.
(2)
(1)
3.2.20.1 Discussion—The procedure employs central or an-
3.2.8 compensator, full-wave (or red plate), n—a compen-
nular stops placed in the objective back focal plane of a
sator (usually a plate of gypsum, selenite or quartz) that
microscope. Using an annular stop with the substage iris
introduces a fixed retardation between 530 to 550 nm (approxi-
closed, a fiber mounted in a high dispersion medium shows a
mately the retardation of the first order red color on the
colored boundary of a wavelength where the fiber and the
Michel-Lévy chart).
medium match in refractive index. Using a central stop, the
(1, 2)
fiber shows colors complementary to those seen with an
3.2.9 compensator, quarter-wave, n—a compensator (usu-
annular stop.
ally a mica plate) that introduces a fixed retardation between
~137–147 nm (approximately the retardation of first-order gray 3.2.21 dye, n—soluble substances that add color to textiles.
on the Michel-Lévy chart). (3)
(1, 2)
3.2.21.1 Discussion—Dyes are classified into groups that
have similar chemical characteristics (for example, aniline,
3.2.10 compensator, quartz wedge, n—a wedge, usually cut
acid, and azo) and also by their method of application (for
from quartz, having continuously variable retardation extend-
example, reactive or direct). They are incorporated into the
ing over several orders (usually 3 to 7) of interference colors.
fiber by chemical reaction, absorption, or dispersion.
(1)
(3)
3.2.11 compensator, Sénarmont, n—a quarter-wave plate
3.2.22 excitation filter, n—a filter used in fluorescence
inserted above the specimen in the parallel “0” position with a
microscopy that transmits specific bands or wavelengths of
calibrated rotating analyzer; measures low retardation and
energy capable of inducing visible fluorescence in various
requires the use of monochromatic light.
substrates.
3.2.12 compensator, tilting (Berek), n—a compensator typi-
cally containing a plate of calcite or quartz, which can be tilted 3.2.23 exclusionary difference, n—a difference in one or
by means of a calibrated drum to introduce incrementally more characteristics between compared items that is sufficient
variable retardation. to determine that the compared items did not originate from the
E2228 − 23a
same source, are not the same source, or do not share the same 3.2.35 medulla, n—the central portion of a hair composed of
composition or classification. a series of discrete cells or an amorphous spongy mass.
3.2.35.1 Discussion—The medulla can be air-filled, and if
3.2.23.1 Discussion—What is sufficient depends on the
so, appears opaque or black using transmitted light or white
performance and limitations of the method used on the material
using reflected light. In animal hair, several types have been
in question.
defined: uniserial or multiserial ladder, cellular or vacuolated,
(4)
and lattice.
3.2.24 extinction, n—the condition in which a birefringent
3.2.36 Michel-Lévy chart, n—a chart relating thickness,
particle appears dark when viewed between crossed polarizers.
birefringence, and retardation so that any one of these variables
(2)
can be determined when the other two are known.
3.2.24.1 Discussion—Most fibers exhibit extinction when
(1)
their long axis is oriented parallel to the privileged direction of
one of the polarizing filters. 3.2.37 microscopical, adj—concerning a microscope or the
use of a microscope.
3.2.25 fluorescence, n—the emission of light by a fiber that
3.2.38 modification ratio, n—a geometrical parameter used
has absorbed light or other electromagnetic radiation of shorter
in the characterization of noncircular fiber cross-sections.
wavelength (higher energy).
3.2.38.1 Discussion—The modification ratio is the ratio in
(2)
size between the outside diameter of the fiber and the diameter
3.2.26 fluorescence microscope, n—a microscope equipped
of the core; it can also be called “aspect ratio.”
with a high energy light source (usually a xenon or mercury
3.2.39 natural fibers, n—a class name for various genera of
vapor lamp) and a set of excitation and barrier filters along with
fibers (including filaments) of: (1) animal (that is, silk and
a dichromatic mirror, used to induce and observe fluorescence
wool); (2) mineral (that is, asbestos); or (3) vegetable origin
in fibers and other particles or materials.
(that is, cotton, flax, jute, and ramie).
3.2.27 inorganic fibers, n—a class of fibers of natural
(3)
mineral origin (for example, chrysotile asbestos) and manufac-
3.2.40 pigment, n—a finely-divided insoluble material used
tured mineral origin (for example, fiberglass).
to deluster or color fibers (for example, titanium dioxide and
3.2.28 interference colors, n—colors produced by the inter-
iron oxide).
ference of two out-of-phase rays of white light when a
(3)
birefringent material is observed at a non-extinction position
3.2.41 plane polarized light, n—light in which the electric
between crossed polars.
field vibrates in one direction in a single plane.
3.2.28.1 Discussion—The retardation at a particular point in
3.2.42 polarized light, n—a bundle of light rays with a
a birefringent fiber can be determined by comparing the
single propagation direction and a single perpendicular vibra-
observed interference color to the Michel-Lévy chart.
tion direction.
3.2.29 isotropic, adj—a characteristic of an object in which
(1)
the refractive index remains constant irrespective of the direc-
3.2.43 polarized light microscope, n—a microscope
tion of propagation or vibration of the light through the object.
equipped with two polarizing filters, one below the stage (the
(1)
polarizer) and one above the stage (the analyzer).
3.2.30 light microscope, n—a microscope that employs light
3.2.43.1 Discussion—When the polarizer and analyzer are
in the visible portion of the electromagnetic spectrum.
inserted into the light path and orientated at 90° to each other,
3.2.31 lignin, n—the majority non-carbohydrate portion of then objects are being observed under crossed polars.
wood; it is an amorphous polymeric substance that cements
3.2.44 privileged direction (of a polarizer), n—the direction
cellulosic fibers together and is the principal constituent of
of vibration to which light emerging from a polarizer has been
woody cell walls.
restricted.
3.2.44.1 Discussion—In modern microscopes, the polariz-
3.2.32 lumen, n—the cavity or central canal present in many
er’s privileged direction is oriented in the east-west direction
natural fibers (for example, cotton, flax, ramie, jute, hemp); its
and the analyzer’s privileged direction is oriented in the
presence and structure are often useful aids in identification.
north-south direction.
3.2.33 luster, n—the gloss or shine possessed by a fiber,
3.2.45 refractive index (n), n—the ratio of the velocity of
resulting from its reflection of light; the luster of manufactured
light in a vacuum to the velocity of light in some medium.
fibers is often modified by use of a delustering pigment.
(1)
3.2.34 manufactured fiber, n—a class name for various
3.2.46 relative refractive index, n—the estimate of the
genera of fibers (including filaments) produced from fiber-
refractive index of a fiber in relation to the index of its
forming substances which can be (1) polymers synthesized
surrounding medium.
from chemical compounds [synthetic fibers], (2) modified or
transformed natural polymers [regenerated fibers], and (3) 3.2.47 retardation (r), n—the actual distance between two
minerals, for example, glasses. doubly refracted rays as they emerge from an anisotropic fiber;
(3) dependent upon the difference in the two refractive indices,
E2228 − 23a
ni2n', and the thickness of the fiber. 6. Sample Handling
3.2.48 sign of elongation, n—a property of fibers referring
6.1 The general handling and tracking of samples should
to the elongation of a fiber in relation to refractive indices.
meet or exceed the requirements of Practice E1492 and Guide
(1) E1459.
3.2.48.1 Discussion—If the fiber is elongated in the direc-
6.2 Items of evidence are visually inspected and forceps can
tion of the higher refractive index, it is said to have a positive
be used to remove fibers of interest. Simple magnifiers and
sign of elongation; if the fiber is elongated in the direction of
stereomicroscopes, with a variety of illumination techniques,
the lower refractive index, it is negative.
can also be employed.
(1)
6.3 Other methods such as tape lifting or gentle scraping are
3.2.49 spherulites, n—spheres composed of needles or rods
usually conducted after a visual examination.
all oriented perpendicular to the outer surface, or a plane
6.3.1 Tape lifts are placed on clear plastic sheets, glass
section through such a sphere; a common form of polymer
microscope slides, or another uncontaminated substrate that
crystallization from melts or concentrated solutions.
eases the search and removal of selected fibers.
(2)
6.3.2 Tape should not be attached to paper or cardboard.
3.2.50 stereomicroscope, n—a microscope containing two
6.3.3 Tapes should not be over loaded.
separate optical paths, one for each eye, giving a three-
6.3.4 Fibers on tape lifts are removed using forceps, other
dimensional view of a specimen.
microscopic tools, or solvents (5-10).
3.2.51 surface dye, n—a colorant bound to the surface of a
6.4 The material recovered is examined with a stereomicro-
fiber.
scope to isolate fibers of interest for further analysis.
3.2.52 synthetic fibers, n—a class of manufactured poly-
6.5 Care should be taken to ensure contamination does not
meric fibers, which are synthesized from chemical compounds
occur.
(for example, nylon and polyester).
6.5.1 Questioned and known items are examined in separate
3.2.53 technical fiber, n—a bundle of natural fibers (for
areas or at different times, or both.
example, hemp, jute, and sisal) composed of individual elon-
6.5.2 The work area and tools are thoroughly cleaned and
gated cells that can be physically or chemically separated and
inspected before examining items that are to be compared.
examined microscopically for identifying characteristics.
7. Performance Verification
3.2.54 thickness (T), n—the optical path through a fiber used
for the calculation of birefringence.
7.1 Equipment—Ideally, the two microscope bases and the
optical bridge of a comparison microscope are provided as a
3.2.55 ultimates, n—individual fibers from a technical fiber
unit by the manufacturer, with the condensers, objectives,
(see 3.2.53).
eyepieces and other optical components matched to each other.
An integrated system allowing delivery of light of the same
4. Significance and Use
intensity and color temperature to both specimens is also
4.1 Microscopical examination is generally a non-
highly desirable. Alternatively, suitable filters (for example,
destructive, rapid, and reproducible means of determining the
color balancing or neutral density filters) can be introduced into
microscopic characteristics, optical properties, and generic
one or both light paths to provide consistent illumination.
polymer type of textile fibers.
Adjustment of lamp rheostats or aperture settings is not
4.2 Side-by-side microscopical comparisons provide a recommended for balancing illumination. If separate illumina-
highly discriminating and efficient method of determining if tion systems are used for the two bases, both bulbs should have
two or more fibers can be differentiated. approximately the same color temperature and always be
replaced at the same time.
4.3 This guideline requires specific pieces of instrumenta-
7.1.1 Effective use of a comparison microscope requires that
tion outlined herein.
the optics and illumination of the two bases be as closely
matched as possible.
5. Summary of Guide
7.2 For uniform illumination, the illumination conditions
5.1 Textile fibers are typically mounted on glass or quartz
are adjusted to those that will be used for sample examination,
microscope slides in a mounting medium under a cover slip.
including proper modified Köhler illumination.
5.2 Fibers are examined microscopically with a combina-
7.3 The balance for light intensity, color temperature, and
tion of various illumination sources, filters, and instrumenta-
overall optical quality should be checked prior to each use of
tion attached to a microscope to determine their polymer type
the microscope and adjusted as necessary. This can be done by
and record any microscopic characteristics and optical proper-
using one or more pairs of test slides made from two sections
ties.
of the same fiber cut in half, with the two halves mounted on
5.3 Known and questioned fibers are compared to determine separate slides. Known red, blue, and green synthetic fiber
if they exhibit the same microscopic characteristics and optical samples should be used to evaluate color balance over the
properties. visible spectrum. Place one test slide on each stage and verify
E2228 − 23a
with side-by-side examination, using each objective, that the staining and the Becke line method (21). Cargille refractive
fiber samples are microscopically indistinguishable. index liquids are suitable for this purpose and are recom-
mended for refractive index measurements of fibers.
7.4 The magnification of corresponding objectives on each
base (for example, 10× versus 10×) should be compared prior 8.3 Optical and Physical Characteristics of Fibers—
to initial use of the microscope. This can be accomplished by Detailed discussions of optical characteristics and their deter-
using a stage micrometer scale and an eyepiece micrometer or mination are provided by McCrone (22-25); McCrone,
an image analysis system. Alternatively, use the test slides in McCrone, and Delly (21); Bloss (26); Chamot and Mason (27);
7.3 to confirm that the fibers do appear to have the same Hartshorne and Stuart (28); and Stoiber and Morse (29).
diameter, and thus the magnification across the system is Polarized light microscopy shall be used to characterize the
consistent. Once uniform magnification for the two bases has optical properties of the fibers.
been verified, it should not need to be repeated unless one or 8.3.1 Observed Color—The color should be observed in
more optical components are replaced or cleaned. transmitted light, with a blue daylight filter or other suitable
color correction in the light path, if needed. It should be noted
8. Analysis
whether fibers are dyed, surface dyed, or pigmented. Variation
in color along the length of individual fibers or between fibers
8.1 Preliminary Examination—Fibers are examined with a
in a sample should also be noted. The use of ultraviolet
stereomicroscope. Physical features such as crimp, length,
(UV)-visible microspectrophotometry is recommended to fur-
color, relative diameter, luster, apparent cross-section, damage,
ther compare the fiber samples.
and adhering debris are assessed. Fibers can then be tentatively
8.3.2 Dichroism—Dichroism can be exhibited by certain
classified into broad groups such as manufactured, natural, or
dyed or pigmented fibers, as well as some mineral fibers.
inorganic. If the sample contains yarns, threads, or sections of
Dichroism is observed by viewing a fiber in plane polarized
fabric, construction should be documented [(11-13) and
light, oriented parallel to the privileged direction of the
AATCC Test Methods 20].
polarizer, then rotating the stage 90 degrees. The substage iris
8.2 Mounting Media—Fibers that are to be microscopically
diaphragm should be opened to a low contrast position for this
examined and compared at higher magnifications are mounted
observation. Any change in color should be noted.
in an appropriate mounting medium. When using a comparison
8.3.3 Refractive Index:
microscope, the same mountant should be used for both
8.3.3.1 The majority of transparent fibers display two prin-
questioned and known fibers. Many suitable media are avail-
cipal refractive indices (that is, they behave as anisotropic,
able as temporary and permanent fiber mounts. The choice of
uniaxial crystals), one for light polarized parallel to the long
mountant depends on availability, the particular application(s),
axis of the fiber (ni) and one for light polarized perpendicular
and examiner preference; however, the following certain cri-
to the long axis of the fiber (n'). For fibers examined in
teria (9, 14-19) should be met:
8.2.1 An examiner should be aware of the possible delete- unpolarized light, a third quantity, n (defined as @2 n '
iso
rious effects that a mounting medium (especially solvent-based
1 n ), can also be estimated. Since refractive index varies
i#
media) can have on textile fibers, particularly when mounted
with wavelength and temperature, a standard refractive index
for a long time. It is preferable that the mounted fibers
(n), is defined for all transparent materials as the refractive
previously examined microscopically be used throughout the
index at a wavelength of 589 nm (the D line of sodium) at
analytical scheme. If fibers are to be removed for further
25 °C.
testing, the mounting medium should be removed with a
8.3.3.2 The refractive indices of a fiber can be determined
solvent that will not alter the fiber.
by several methods. Whatever the method used, determination
8.2.2 If a solvent-based mounting medium is used for
of ni and n' should be made using plane polarized light with
refractive index determination, the index of the mountant
the fiber aligned parallel and perpendicular to the privileged
should be checked periodically against solid refractive index
direction of the polarizer, respectively. The vibration direction
standards and, if necessary, readjusted to its proper value by the
of the polarizer should coincide with the horizontal line of the
addition of solvent (20). Additionally, the refractive index of
eyepiece graticule.
the medium can be measured directly (that is, by using an Abbe
refractometer) and the value recorded by the examiner. If such 8.3.3.3 Refractive index measurements can be relative or
a medium is used for permanent mounts, the examiner should exact.
be aware of the different refractive indices for the fluid medium (1) A relative refractive index measurement involves: (1)
and the resin after solvent evaporation. determining whether an immersed object is higher or lower in
8.2.3 The tolerance at n shall be known for liquids used for refractive index than the immersion medium using the Becke
D
refractive index determinations of fibers. For most refractive line method, and (2) estimating the approximate refractive
index liquids, this value is determined by the manufacturer. index based upon the amount of contrast between the fiber and
Alternatively, the refractive index values can be measured the medium. The degree of contrast shows the amount of
using an Abbe refractometer. To make appropriate temperature refractive index difference between the fiber and the medium.
corrections, values for the temperature coefficient (dn/dt) for (2) Numerical values for ni and n' of a fiber can be
each liquid should be available, as well as a thermometer determined by immersing the fiber or fibers in successive
covering the range 20 to 30 °C, calibrated in tenths of a degree. liquids and observing with a filter (for example, sodium D at
High dispersion liquids (V < 30) are desirable for dispersion 25°C) until the minimum contrast between the specimen and
E2228 − 23a
the mounting medium is achieved at the particular orientation chart. Take care when interpreting results from deeply dyed
relative to the polarizer. Refractive indices can also be deter- fibers, as the dye can obscure the interference colors. A wedge
mined by dispersion staining. slice through the fiber or the use of various compensators, such
as the Sénarmont, quartz wedge, and tilting (Berek), can be
8.3.3.4 Dispersion Staining—Dispersion staining is an alter-
used to make a more accurate determination of retardation
native to the Becke line method for refractive index determi-
(32). When measuring retardation of a fiber using a tilting
nation. It is particularly useful for the identification of asbestos
compensator or quartz wedge, ensure no error has been
fibers, but can also be applied to the identification of other fiber
introduced due to differences in dispersion of birefringence
types (1, 21, 30, 31).
between the compensator and the fiber. This is of special
(1) Dispersion staining is performed using an objective that
concern with the examination of fibers with high birefringence.
employs opaque central or annular stops placed in the back
The birefringence of noncircular fibers can be estimated by
focal plane. Special objectives of this type can be purchased
measuring both retardation and thickness at two points along
commercially or prepared in the laboratory by introducing
the fiber that represent their highest and lowest values (33). The
stops into the back focal plane of a normal objective (usually
thickness can also be measured using a cross-section of the
10× or 20×). Using an annular stop with the substage iris
fiber.
closed, a fiber or other particle shows a colored boundary of a
8.3.5 Sign of Elongation—For a birefringent fiber, the sign
wavelength where the fiber and the medium match in refractive
of elongation is positive (+) if ni.n' and negative (–) if ni
index. Using a central stop, the fiber shows colors complemen-
,n'. The common manufactured fibers with a birefringence
tary to those seen with an annular stop. Central stop observa-
higher than 0.010 have a positive sign of elongation. Full or
tion (in which particles have colored borders against a black
quarter wave compensators are commonly used to make this
background) is more commonly employed.
determination for fibers with birefringence less than 0.010,
(2) For optimum use of dispersion staining, mounting
which exhibit first order gray or white retardation colors (9,
media with a high dispersion should be used. Cargille high
26). To determine sign of elongation for a low birefringence
dispersion refractive index liquids are recommended. Carefully
fiber, the fiber is oriented perpendicular to the orientation of the
clean slides and cover slips of dirt, debris, and finger marks.
compensator between crossed polars. A full wave (first order
When using a central stop, center the stop in the back focal
red) compensator, for example, is inserted with the slow
plane and ensure that it is large enough to block direct light
direction (Z direction on the compensator) parallel to the length
rays from a fully closed or almost fully closed substage iris
of the fiber. Fibers with a positive sign of elongation appear
diaphragm. With the dispersion staining objective focused on a
blue (higher interference color relative to first order red) in this
specimen, the suitable size and centration of the stop can be
orientation, while fibers with a negative sign of elongation
verified by inserting the Bertrand lens and observing the back
appear orange (lower interference color).
focal plane.
(3) To observe dispersion staining colors, focus the disper-
8.3.6 Diameter—The diameter of circular fibers can be
sion staining objective on a fiber in plane polarized light (single
measured using an eyepiece micrometer or an image analysis
polarizer) and orient the fiber in an ni or n' direction relative
system, calibrated with a micrometer slide for each microscope
to the polarizer. Close the substage iris until a dark background
objective or magnification. Noncircular fibers require special
is obtained and observe the color bordering the fiber. Rotate the
considerations (34). If fiber diameters are not uniform within a
stage 90 degrees to observe the color for the other index. Based
sample, or if different aspects are presented by non-circular
on the dispersion staining colors observed, the matching
fibers, a determination of the range of diameters exhibited by
wavelengths for the specimen and the liquid can be determined
the sample is recommended. Measurements should be made at
by reference to published tables or color charts and the
the highest magnification that is practical, with the substage iris
refractive indices of the specimen relative to the liquid can be
opened to a position of low to moderate contrast, so that the
estimated.
edges of the fiber are defined, but not too dark.
(4) By mounting a fiber in a series of liquids and observing
8.3.7 Cross-Section—When viewed longitudinally on glass
dispersion staining colors for each, dispersion curves for the ni
slides in a suitable mountant, the apparent cross-sectional
and n' refractive indices of a fiber can be plotted, and the
shape of fibers can often be determined by slowly focusing
indices at 589 nm determined more precisely.
through the fiber (optical sectioning) or by observing the
8.3.4 Birefringence—For a fiber displaying two refractive
different interference colors and their relative positions across
indices, birefringence is defined as |ni2n'|. Birefringence is the width of the fiber. Actual fiber cross-sections provide the
determined by measuring ni and n' and using the above
best information on cross-sectional shape. Manufactured and
equation or by determining the retardation with the correspond- vegetable fibers can be cross-sectioned anywhere on their
ing thickness of the fiber and calculated with the following
length (35-41). Animal hairs can be cross-sectioned to yield
equation: additional identifying characteristics (42, 43). When observing
manufactured fiber cross-sections, the general shape, distribu-
Retardation ~nm!
tion of delustrant, or pigment particles, or combination thereof;
Birefringence 5 (1)
1000 × Thickness ~µm!
the presence and size of spherulites or voids; depth of dye
8.3.4.1 The retardation can be estimated by observing the penetration; and surface treatments should be recorded when
interference color displayed at the point where the thickness of present. Cross-sectioning is also useful in the recognition and
the fiber is measured and comparing it to the Michel-Lévy examination of bicomponent fibers. The fiber dimensions
E2228 − 23a
measured from a cross section can be used for the calculation temperature(s), should be recorded. Changes in the physical
of birefringence and the determination of the modification ratio state of a fiber are often indicated by changes in birefringence.
of multi-lobed fibers. Since manufactured fibers are composed of mixtures of chemi-
cal compounds rather than pure polymers and are a combina-
8.3.8 Modification Ratio—The modification ratio of non-
tion of crystalline and amorphous regions, changes are nor-
circular fibers can be calculated by obtaining an image of the
mally observed over a temperature range rather than at a single
fiber cross-section, and using a circle template or image
melting point (9, 11, 38, 40, 51-55). Fibers should be mounted
analysis system to determine the sizes of the circumscribing
and inscribing circles for that shape. The modification ratio is in an inert, heat-resistant medium, such as high-temperature
stable silicone oil, to ensure reproducible melting behavior (54,
the ratio of the larger circle’s diameter to the smaller circle’s
diameter. This value can help to identify a particular manufac- 56). Accurate and reproducible results are best obtained using
a heating rate of no greater than 1 to 2 °C ⁄min when near the
turer or end use of a fiber.
initial melting temperature. The hot stage should be calibrated
8.3.9 Delustrant, Pigment, and Filler—The presence or
using appropriate standards, following established guidelines
absence of delustrant, pigment, and filler particles, as well as
(57). The recommended melting point apparatus should be
their size, shape, distribution, abundance, and general
adjustable for temperatures from ambient to at least 300 °C, in
appearance, are useful comparative features. Also, the presence
increments of 0.1 °C, and should allow a heating rate of as low
of these particles shows conclusively that a fiber is
as 1 °C ⁄min (56-65).
manufactured, rather than natural. While not indicative of any
particular generic fiber type, these particles can be character- 8.4.3 Scanning Electron Microscopy—Scanning electron
istic of end use properties needed by a manufacturer, such as
microscopy with energy dispersive X-ray spectroscopy (SEM-
antimony trioxide particles being indicative of fire retardant EDS) is used as an imaging and microanalytical tool in the
material.
characterization of fibers (66). Fiber surface morphology can
be examined with great depth of field at continually variable
8.3.10 Surface Characteristics—Fiber surface
magnifications. Fibers and prepared cross sections mounted on
characteristics, such as manufacturing striations, damage, and
specimen stubs can be conductively coated to prevent possible
surface debris (that is, blood or other foreign material) should
electron beam charging. The use of a suitable calibration
be described. Surface striations are more apparent in a mount-
standard is recommended for the accurate measurement of fiber
ing medium of refractive index significantly different from
cross sections.
those of the fiber.
8.3.11 Fluorescence—Fluorescence can arise from fibers
8.4.3.1 Applications of SEM-EDS to fiber analysis include
themselves, dyes, other additives from the finishing process, the characterization of fiber cross sections, identification of
laundering, chemical treatment/damage, as well as surface
pigments, delustrants, and the presence of nanoparticles by
debris. Fibers should be mounted in a low- to non-fluorescent elemental analysis, fiber damage due to cuts and tears, trace
medium to best observe fluorescence. Examination using
debris on fibers, and surface feature modifications such as
various combinations of excitation and barrier filters is desir- washer/dryer abrasion and acid washed treatment of denim
able. At each excitation wavelength, the color and intensity or garments. Authors have examined fiber bonding in nonwoven
absence of fluorescence emission should be noted (9, 11, fabrics and shrink-proofing treatment of wool have also been
44-48). studied. Surface imaging using the SEM as an aid in the
identification of animal hair scale structure has also been
8.4 Additional Characterization Techniques:
reported (41, 67-73).
8.4.1 Solubility—Solubility testing can provide supplemen-
tal information to optical methods of characterization, but since
9. Classification and Identification
it is a destructive method, it should be used only when
sufficient sample is available and non-destructive methods
9.1 Manufactured Fibers—After preliminary examination
have been exhausted. Possible reactions of fibers to solvents
and general classification by use of a stereomicroscope, the
include partial and complete solubility, swelling, shrinking,
generic fiber type can usually be identified using a polarized
gelling, and color change. If solubility tests are used as part of light microscope. Manufactured fiber types are best identified
an identification scheme, appropriate controls should be run
by determining optical properties such as refractive indices,
following the laboratory’s quality assurance and quality control
birefringence, and sign of elongation. Solubility and melting
(QA/QC) guidelines for a lot or batch of reagents or solvents.
point determination, while not recommended as primary meth-
It is desirable to view known and questioned fibers simultane-
ods of identification, can assist in confirming the generic type
ously under a microscope when comparing their solubilities
(for example, nylon) and in identifying sub-groups within
[(9, 49, 50) and Test Methods D276].
particular generic types (for example, nylon 6,6; nylon 12).
8.4.2 Hot Stage Microscopy—A polarized light microscope Fourier transform infrared spectroscopy (FTIR) is recom-
equipped with a hot stage is recommended for observations of mended to identify sub-groups within synthetic fiber types.
the effect of heat on fibers. Since it is a destructive method, it Elemental analysis by SEM-EDS analysis is useful in sub-
should be used only when sufficient sample is available and typing glass fibers, as is refractive index measurement. Physi-
non-destructive methods have been exhausted. Using slightly cal features such as diameter, cross-section, modification ratio
uncrossed polars, droplet formation, contraction, softening, and surface treatment, while not necessarily characteristic of a
charring, and melting of fibers over a range of temperatures can particular fiber type, can aid in identifying or eliminating
be observed; these observations, including melting possible end uses (for example, trilobal carpet fibers) and are
E2228 − 23a
also important comparative features. Features such as color, teased apart, and paper re-pulped for the examination of
dichroism, delustering, and fluorescence are primarily of use individual cells. Relative thickness of cell walls and lumen, cell
for comparison of different fiber samples. length and diameter, cell end shape, and the presence, type, and
distribution of dislocations should be noted. Staining tests
9.2 Glass Fibers—Glass fibers are encountered in building
using Herzberg or Graff’s C-stain can also be useful for
materials, insulation products and in fabrics. Glass fibers are
identification purposes. The direction of twist of the cellulose
also called manufactured vitreous fibers (74). Based on the
in the cell wall can also be determined using the Herzog test
starting materials used to produce glass fibers, they can be
(81). Other characteristic cells should be noted and compared
placed into three categories: fiberglass (continuous and non-
to authentic specimens (82-84). Wood pulp fibers from paper or
continuous), mineral wool (rock wool and slag wool), and
cardboard can be distinguished from other cellulosic fibers by
refractory ceramic fibers (glass ceramic fibers). Single crystal
morphology or staining tests. Distinction between hardwood
and polycrystalline refractory fibers such as aluminum oxide,
and softwood and more specific genus or species identification
silicon carbide, zirconium oxide, and carbon are not included
can also be possible for wood fibers (81, 83-86).
because they are not considered glass fibers.
9.3.3 Mineral Fibers—Natural mineral fibers are commonly
9.2.1 Analysis and Characterization of Glass Fibers—Glass
called asbestos, which is a general term for many naturally-
fibers are normally identified by their morphology and isotro-
occurring fibrous, hydrated silicate minerals. The asbestos
pic nature. The presence or absence of coating resins and of
minerals include chrysotile, amosite, crocidolite, fibrous
spheres and slugs can indicate an end use (for example,
tremolite/actinolite, and fibrous anthophyllite. Chrysotile be-
building insulation) and are also useful comparative features.
longs to the serpentine group of minerals that are layer
Light microscopy, together with classical immersion methods,
silicates. The other asbestos minerals are amphiboles and are
can be used to determine the refractive index for the classifi-
classified as chain silicates. Asbestos fibers alone, or mixed
cation and comparison of glass fibers. The dispersion staining
with other components, can occur in building materials and
technique can be used when determining the refractive index
insulation products. Chrysotile is the only asbestos mineral that
and variation of the refractive index within a sample. SEM-
would be encountered as a woven fabric, but any of the
EDS can be used to provide elemental composition for
asbestos minerals can be found in pressed sheets such as
purposes of classification and comparison.
gaskets. Take care when analyzing asbestos fibers because they
9.3 Natural Fibers—Most natural fiber types are best iden-
are considered a potential health hazard.
tified by their physical and morphological characteristics.
9.3.3.1 Asbestos minerals can be easily identified by their
Optical properties such as refractive indices and birefringence
optical properties using polarized light microscopy and the
are of more limited use in identifying or comparing natural
dispersion staining technique (1, 30). Scanning electron mi-
fibers than in the analysis of manufactured fibers, with the
croscopy or transmission electron spectroscopy with energy
exception of identifying specific types of asbestos. For natural
dispersive spectrometry can also be used to characterize the
fiber comparisons, color is the
...