NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
ASBESTOS and OTHER FIBERS by PCM 7400
FORMULA: Various MW: Various CAS: see Synonyms RTECS: Various
METHOD: 7400, Issue 2 EVALUATION: FULL Issue 1: Rev. 3 on 15 May 1989
Issue 2: 15 August 1994
OSHA: 0.1 asbestos ber (> 5 µm long)/cc; 1 f/cc, 30 min
excursion; carcinogen
MSHA: 2 asbestos bers/cc
NIOSH: 0.1 f/cc (bers > 5 µm long), 400 L; carcinogen
ACGIH: 0.2 f/cc crocidolite; 0.5 f/cc amosite; 2 f/cc chrysotile
and other asbestos; carcinogen
PROPERTIES: solid, brous, crystalline, anisotropic
SYNONYMS [CAS #]: actinolite [77536-66-4] or ferroactinolite [15669-07-5]; amosite [12172-73-5]; anthophyllite [77536-
67-5]; chrysotile [12001-29-5]; serpentine [18786-24-8]; crocidolite [12001-28-4]; tremolite [77536-68-6];
amphibole asbestos [1332-21-4]; refractory ceramic bers [142844-00-6]; brous glass
SAMPLING
SAMPLER: FILTER
(0.45- to 1.2-µm cellulose ester membrane,
25-mm; conductive cowl on cassette)
FLOW RATE*: 0.5 to 16 L/min
VOL-MIN*: 400 L @ 0.1 ber/cc
-MAX*: (step 4, sampling)
*Adjust to give 100 to 1300 ber/mm²
SHIPMENT: routine (pack to reduce shock)
SAMPLE
STABILITY: stable
BLANKS: 2 to 10 eld blanks per set
ACCURACY
RANGE STUDIED: 80 to 100 bers counted
BIAS: see EVALUATION OF METHOD
OVERALL PRECISION (
): 0.115 to 0.13 [1]
ACCURACY: see EVALUATION OF METHOD
MEASUREMENT
TECHNIQUE: LIGHT MICROSCOPY, PHASE CONTRAST
ANALYTE: bers (manual count)
SAMPLE
PREPARATION: acetone - collapse/triacetin - immersion
method [2]
COUNTING
RULES: described in previous version of this
method as “A” rules [1,3]
EQUIPMENT: 1. positive phase-contrast microscope
2. Walton-Beckett graticule (100-µm eld
of view) Type G-22
3. phase-shift test slide (HSE/NPL)
CALIBRATION: HSE/NPL test slide
RANGE: 100 to 1300 bers/mm² lter area
ESTIMATED LOD: 7 bers/mm² lter area
PRECISION (
): 0.10 to 0.12 [1]; see EVALUATION OF
METHOD
APPLICABILITY: The quantitative working range is 0.04 to 0.5 ber/cc for a 1000-L air sample. The LOD depends on sample
volume and quantity of interfering dust, and is <0.01 ber/cc for atmospheres free of interferences. The method gives an
index of airborne bers. It is primarily used for estimating asbestos concentrations, though PCM does not dierentiate
between asbestos and other bers. Use this method in conjunction with electron microscopy (e.g., Method 7402) for assis-
tance in identication of bers. Fibers < ca. 0.25 µm diameter will not be detected by this method [4]. This method may be
used for other materials such as brous glass by using alternate counting rules (see Appendix C).
INTERFERENCES: If the method is used to detect a specic type of ber, any other airborne ber may interfere since all
particles meeting the counting criteria are counted. Chain-like particles may appear brous. High levels of non-brous dust
particles may obscure bers in the eld of view and increase the detection limit.
OTHER METHODS: This revision replaces Method 7400, Revision #3 (dated 5/15/89).
NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
ASBESTOS and OTHER FIBERS by PCM: METHOD 7400, Issue 2, dated 15 August 1994 - Page 2 of 15
REAGENTS:
1. Acetone,* reagent grade.
2. Triacetin (glycerol triacetate), reagent grade.
*See SPECIAL PRECAUTIONS.
EQUIPMENT:
1. Sampler: eld monitor, 25-mm, three-piece
cassette with ca. 50-mm electrically conductive
extension cowl and cellulose ester lter, 0.45-
to 1.2-µm pore size, and backup pad.
NOTE 1: Analyze representative lters for ber
background before use to check for
clarity and background. Discard the
lter lot if mean is ≥ 5 bers per 100
graticule elds. These are dened
as laboratory blanks. Manufacturer-
provided quality assurance checks on
lter blanks are normally adequate as
long as eld blanks are analyzed as
described below.
NOTE 2: The electrically conductive extension
cowl reduces electrostatic eects.
Ground the cowl when possible
during sampling.
NOTE 3: Use 0.8-µm pore size lters for
personal sampling. The 0.45-µm
lters are recommended for sampling
when performing TEM analysis on the
same samples. However, their higher
pressure drop precludes their use with
personal sampling pumps.
NOTE 4: Other cassettes have been proposed
that exhibit improved uniformity of
ber deposit on the lter surface, e.g.,
bellmouthed sampler (Envirometrics,
Charleston, SC). These may be
used if shown to give measured
concentrations equivalent to sampler
indicated above for the application.
2. Personal sampling pump, battery or line-
powered vacuum, of sucient capacity to
meet ow-rate requirements (see step 4 for
ow rate), with exible connecting tubing.
3. Wire, multi-stranded, 22-gauge; 1″ hose clamp
to attach wire to cassette.
4. Tape, shrink- or adhesive-.
5. Slides, glass, frosted-end, pre-cleaned, 25- ×
75-mm.
6. Cover slips, 22- × 22-mm, No. 1½, unless
otherwise specied by microscope
manufacturer.
7. Lacquer or nail polish.
8. Knife, #10 surgical steel, curved blade.
9. Tweezers.
NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
ASBESTOS and OTHER FIBERS by PCM: METHOD 7400, Issue 2, dated 15 August 1994 - Page 3 of 15
EQUIPMENT (continued):
10. Acetone ash vaporization system for
clearing lters on glass slides (see ref. [5]
for specications or see manufacturer’s
instructions for equivalent devices).
11. Micropipets or syringes, 5-µL and 100- to
500-µL.
12. Microscope, positive phase (dark) contrast,
with green or blue lter, adjustable eld
iris, 8 to 10× eyepiece, and 40 to 45× phase
objective (total magnication ca. 400×);
numerical aperture = 0.65 to 0.75.
13. Graticule, Walton-Beckett type with 100-µm
diameter circular eld (area = 0.00785 mm²)
at the specimen plane (Type G-22). Available
from Optometrics USA, P.O. Box 699, Ayer, MA
01432 [phone (508)-772-1700], and McCrone
Accessories and Components, 850 Pasquinelli
Drive, Westmont, IL 60559 [phone (312)
887-7100].
NOTE: The graticule is custom-made for each
microscope. (see APPENDIX A for the
custom-ordering procedure).
14. HSE/NPL phase contrast test slide, Mark II.
Available from Optometrics USA (address
above).
15. Telescope, ocular phase-ring centering.
16. Stage micrometer (0.01-mm divisions).
SPECIAL PRECAUTIONS: Acetone is extremely ammable. Take precautions not to ignite it. Heating
of acetone in volumes greater than 1 mL must be done in a ventilated laboratory fume hood using a
ameless, spark-free heat source.
SAMPLING:
1. Calibrate each personal sampling pump with a representative sampler in line.
2. To reduce contamination and to hold the cassette tightly together, seal the crease between the
cassette base and the cowl with a shrink band or light colored adhesive tape. For personal sampling,
fasten the (uncapped) open-face cassette to the worker’s lapel. The open face should be oriented
downward.
NOTE: The cowl should be electrically grounded during area sampling, especially under conditions
of low relative humidity. Use a hose clamp to secure one end of the wire (Equipment, Item 3)
to the monitor’s cowl. Connect the other end to an earth ground (i.e., cold water pipe).
3. Submit at least two eld blanks (or 10% of the total samples, whichever is greater) for each set of
samples. Handle eld blanks in a manner representative of actual handling of associated samples in
the set. Open eld blank cassettes at the same time as other cassettes just prior to sampling. Store
top covers and cassettes in a clean area (e.g., a closed bag or box) with the top covers from the
sampling cassettes during the sampling period.
4. Sample at 0.5 L/min or greater [6]. Adjust sampling ow rate, (L/min), and time, t (min), to produce
a ber density,
, of 100 to 1300 bers/mm² (3.85×10⁴ to 5×10⁵ bers per 25-mm lter with eective
NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
ASBESTOS and OTHER FIBERS by PCM: METHOD 7400, Issue 2, dated 15 August 1994 - Page 4 of 15
collection area = 385 mm²) for optimum accuracy. These variables are related to the action level
(one-half the current standard), (bers/cc), of the brous aerosol being sampled by:
.
NOTE 1: The purpose of adjusting sampling times is to obtain optimum ber loading on the lter.
The collection eciency does not appear to be a function of ow rate in the range of 0.5
to 16 L/min for asbestos bers [7]. Relatively large diameter bers (>3 µm) may exhibit
signicant aspiration loss and inlet deposition. A sampling rate of 1 to 4 L/min for 8 h is
appropriate in atmospheres containing ca. 0.1 ber/cc in the absence of signicant amounts
of non-asbestos dust. Dusty atmospheres require smaller sample volumes (≤400 L) to obtain
countable samples. In such cases take short, consecutive samples and average the results
over the total collection time. For documenting episodic exposures, use high ow rates (7
to 16 L/min) over shorter sampling times. In relatively clean atmospheres, where targeted
ber concentrations are much less than 0.1 ber/cc, use larger sample volumes (3000 to
10000 L) to achieve quantiable loadings. Take care, however, not to overload the lter with
background dust. If ≥50% of the lter surface is covered with particles, the lter may be too
overloaded to count and will bias the measured ber concentration.
NOTE 2: OSHA regulations specify a minimum sampling volume of 48 L for an excursion
measurement, and a maximum sampling rate of 2.5 L/min [3].
5. At the end of sampling, replace top cover and end plugs.
6. Ship samples with conductive cowl attached in a rigid container with packing material to prevent
jostling or damage.
NOTE: Do not use untreated polystyrene foam in shipping container because electrostatic forces
may cause ber loss from sample lter.
SAMPLE PREPARATION:
NOTE 1: The object is to produce samples with a smooth (non-grainy) background in a medium with
refractive index ≤ 1.46. This method collapses the lter for easier focusing and produces
permanent (1–10 years) mounts which are useful for quality control and interlaboratory
comparison. The aluminum “hot block” or similar ash vaporization techniques may be
used outside the laboratory [2]. Other mounting techniques meeting the above criteria
may also be used (e.g., the laboratory fume hood procedure for generating acetone vapor
as described in Method 7400—revision of 5/15/85, or the non-permanent eld mounting
technique used in P&CAM 239 [3,7–9]). Unless the eective ltration area is known,
determine the area and record the information referenced against the sample ID number
[1,9–11].
NOTE 2: Excessive water in the acetone may slow the clearing of the lter, causing material to be
washed o the surface of the lter. Also, lters that have been exposed to high humidities
prior to clearing may have a grainy background.
7. Ensure that the glass slides and cover slips are free of dust and bers.
8. Adjust the rheostat to heat the “hot block” to ca. 70 °C [2].
NOTE: If the “hot block” is not used in a fume hood, it must rest on a ceramic plate and be isolated
from any surface susceptible to heat damage.
9. Mount a wedge cut from the sample lter on a clean glass slide.
a. Cut wedges of ca. 25% of the lter area with a curved-blade surgical steel knife using a rocking
motion to prevent tearing. Place wedge, dust side up, on slide.
NOTE: Static electricity will usually keep the wedge on the slide.
b. Insert slide with wedge into the receiving slot at base of “hot block”. Immediately place tip of
a micropipet containing ca. 250 µL acetone (use the minimum volume needed to consistently
clear the lter sections) into the inlet port of the PTFE cap on top of the “hot block” and inject the
NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
ASBESTOS and OTHER FIBERS by PCM: METHOD 7400, Issue 2, dated 15 August 1994 - Page 5 of 15
acetone into the vaporization chamber with a slow, steady pressure on the plunger button while
holding pipet rmly in place. After waiting 3 to 5 s for the lter to clear, remove pipet and slide
from their ports.
CAUTION: Although the volume of acetone used is small, use safety precautions. Work in a
well-ventilated area (e.g., laboratory fume hood). Take care not to ignite the acetone.
Continuous use of this device in an unventilated space may produce explosive acetone
vapor concentrations.
c. Using the 5-µL micropipet, immediately place 3.0 to 3.5 µL triacetin on the wedge. Gently lower
a clean cover slip onto the wedge at a slight angle to reduce bubble formation. Avoid excess
pressure and movement of the cover glass.
NOTE: If too many bubbles form or the amount of triacetin is insucient, the cover slip may
become detached within a few hours. If excessive triacetin remains at the edge of the lter
under the cover slip, ber migration may occur.
d. Mark the outline of the lter segment with a glass marking pen to aid in microscopic evaluation.
e. Glue the edges of the cover slip to the slide using lacquer or nail polish [12]. Counting may
proceed immediately after clearing and mounting are completed.
NOTE: If clearing is slow, warm the slide on a hotplate (surface temperature 50 °C) for up to 15
min to hasten clearing. Heat carefully to prevent gas bubble formation.
CALIBRATION AND QUALITY CONTROL:
10. Microscope adjustments. Follow the manufacturer’s instructions. At least once daily use the
telescope ocular (or Bertrand lens, for some microscopes) supplied by the manufacturer to ensure
that the phase rings (annular diaphragm and phase-shifting elements) are concentric. With each
microscope, keep a logbook in which to record the dates of microscope cleanings and major
servicing.
a. Each time a sample is examined, do the following:
(1) Adjust the light source for even illumination across the eld of view at the condenser iris. Use
Kohler illumination, if available. With some microscopes, the illumination may have to be set
up with bright eld optics rather than phase contract optics.
(2) Focus on the particulate material to be examined.
(3) Make sure that the eld iris is in focus, centered on the sample, and open only enough to fully
illuminate the eld of view.
b. Check the phase-shift detection limit of the microscope periodically for each analyst/microscope
combination:
(1) Center the HSE/NPL phase-contrast test slide under the phase objective.
(2) Bring the blocks of grooved lines into focus in the graticule area.
NOTE: The slide contains seven blocks of grooves (ca. 20 grooves per block) in descending
order of visibility. For asbestos counting, the microscope optics must completely
resolve the grooved lines in block 3 although they may appear somewhat faint, and
the grooved lines in blocks 6 and 7 must be invisible when centered in the graticule
area. Blocks 4 and 5 must be at least partially visible but may vary slightly in visibility
between microscopes. A microscope which fails to meet these requirements has
resolution either too low or too high for ber counting.
(3) If image quality deteriorates, clean the microscope optics. If the problem persists, consult the
microscope manufacturer.
11. Document the laboratory’s precision for each counter for replicate ber counts.
a. Maintain as part of the laboratory quality assurance program a set of reference slides to be
used on a daily basis [13]. These slides should consist of lter preparations including a range of
loadings and background dust levels from a variety of sources including both eld and reference
samples (e.g., PAT, AAR, commercial samples). The Quality Assurance Ocer should maintain
custody of the reference slides and should supply each counter with a minimum of one reference
NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
ASBESTOS and OTHER FIBERS by PCM: METHOD 7400, Issue 2, dated 15 August 1994 - Page 6 of 15
slide per workday. Change the labels on the reference slides periodically so that the counter does
not become familiar with the samples.
b. From blind repeat counts on reference slides, estimate the laboratory intra- and intercounter
precision. Obtain separate values of relative standard deviation (
) for each sample matrix
analyzed in each of the following ranges: 5 to 20 bers in 100 graticule elds, >20 to 50 bers in
100 graticule elds, and >50 to 100 bers in 100 graticule elds. Maintain control charts for each
of these data les.
NOTE: Certain sample matrices (e.g., asbestos cement) have been shown to give poor precision
[9].
12. Prepare and count eld blanks along with the eld samples. Report counts on each eld blank.
NOTE 1: The identity of blank lters should be unknown to the counter until all counts have been
completed.
NOTE 2: If a eld blank yields greater than 7 bers per 100 graticule elds, report possible
contamination of the samples.
13. Perform blind recounts by the same counter on 10% of lters counted (slides relabeled by a person
other than the counter). Use the following test to determine whether a pair of counts by the same
counter on the same lter should be rejected because of possible bias: Discard the sample if the
absolute value of the dierence between the square roots of the two counts (in ber/mm²) exceeds
where = average of the square roots of the two ber counts (in ber/mm²) and
where is the intracounter relative standard deviation for the appropriate count range (in bers)
determined in step 11. For more complete discussions see reference [13].
NOTE 1: Since ber counting is the measurement of randomly placed bers which may be described
by a Poisson distribution, a square root transformation of the ber count data will result in
approximately normally distributed data [13].
NOTE 2: If a pair of counts is rejected by this test, recount the remaining samples in the set and test
the new counts against the rst counts. Discard all rejected paired counts. It is not necessary
to use this statistic on blank counts.
14. The analyst is a critical part of this analytical procedure. Care must be taken to provide a non-
stressful and comfortable environment for ber counting. An ergonomically designed chair should
be used, with the microscope eyepiece situated at a comfortable height for viewing. External
lighting should be set at a level similar to the illumination level in the microscope to reduce eye
fatigue. In addition, counters should take 10- to 20-minute breaks from the microscope every one or
two hours to limit fatigue [14]. During these breaks, both eye and upper back/neck exercises should
be performed to relieve strain.
15. All laboratories engaged in asbestos counting should participate in a prociency testing program
such as the AIHA-NIOSH Prociency Analytical Testing (PAT) Program for asbestos and routinely
exchange eld samples with other laboratories to compare performance of counters.
MEASUREMENT:
16. Center the slide on the stage of the calibrated microscope under the objective lens. Focus the
microscope on the plane of the lter.
17. Adjust the microscope (Step 10).
NOTE: Calibration with the HSE/NPL test slide determines the minimum detectable ber diameter
(ca. 0.25 µm) [4].
18. Counting rules: (same as P&CAM 239 rules [1,10,11]: see examples in APPENDIX B).
a. Count any ber longer than 5 µm which lies entirely within the graticule area.
(1) Count only bers longer than 5 µm. Measure length of curved bers along the curve.
(2) Count only bers with a length-to-width ratio equal to or greater than 3:1.
b. For bers which cross the boundary of the graticule eld:
(1) Count as ½ ber any ber with only one end lying within the graticule area, provided that the
ber meets the criteria of rule a above.
NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
ASBESTOS and OTHER FIBERS by PCM: METHOD 7400, Issue 2, dated 15 August 1994 - Page 7 of 15
(2) Do not count any ber which crosses the graticule boundary more than once.
(3) Reject and do not count all other bers.
c. Count bundles of bers as one ber unless individual bers can be identied by observing both
ends of a ber.
d. Count enough graticule elds to yield 100 bers. Count a minimum of 20 elds. Stop at 100
graticule elds regardless of count.
19. Start counting from the tip of the lter wedge and progress along a radial line to the outer edge.
Shift up or down on the lter, and continue in the reverse direction. Select graticule elds randomly
by looking away from the eyepiece briey while advancing the mechanical stage. Ensure that, as a
minimum, each analysis covers one radial line from the lter center to the outer edge of the lter.
When an agglomerate or bubble covers ca. 1/6 or more of the graticule eld, reject the graticule
eld and select another. Do not report rejected graticule elds in the total number counted.
NOTE 1: When counting a graticule eld, continuously scan a range of focal planes by moving the
ne focus knob to detect very ne bers which have become embedded in the lter. The
small-diameter bers will be very faint but are an important contribution to the total count.
A minimum counting time of 15 s per eld is appropriate for accurate counting.
NOTE 2: This method does not allow for dierentiation of bers based on morphology. Although
some experienced counters are capable of selectively counting only bers which appear to
be asbestiform, there is presently no accepted method for ensuring uniformity of judgment
between laboratories. It is, therefore, incumbent upon all laboratories using this method
to report total ber counts. If serious contamination from non-asbestos bers occurs in
samples, other techniques such as transmission electron microscopy must be used to
identify the asbestos ber fraction present in the sample (see NIOSH Method 7402). In some
cases (i.e., for bers with diameters >1 µm), polarized light microscopy (as in NIOSH Method
7403) may be used to identify and eliminate interfering non-crystalline bers [15].
NOTE 3: Do not count at edges where lter was cut. Move in at least 1 mm from the edge.
NOTE 4: Under certain conditions, electrostatic charge may aect the sampling of bers. These
electrostatic eects are most likely to occur when the relative humidity is low (below 20%),
and when sampling is performed near the source of aerosol. The result is that deposition of
bers on the lter is reduced, especially near the edge of the lter. If such a pattern is noted
during ber counting, choose elds as close to the center of the lter as possible [5].
NOTE 5: Counts are to be recorded on a data sheet that provides, as a minimum, spaces on which to
record the counts for each eld, lter identication number, analyst’s name, date, total bers
counted, total elds counted, average count, ber density, and commentary. Average count
is calculated by dividing the total ber count by the number of elds observed. Fiber density
(bers/mm²) is dened as the average count (bers/eld) divided by the eld (graticule) area
(mm²/eld).
CALCULATIONS AND REPORTING OF RESULTS
20. Calculate and report ber density on the lter, (bers/mm²), by dividing the average ber count
per graticule eld,
, minus the mean eld blank count per graticule eld, , by the graticule
eld area, (approx. 0.00785 mm²):
, bers/mm².
NOTE: Fiber counts above 1300 bers/mm² and ber counts from samples with >50% of lter area
covered with particulate should be reported as “uncountable” or “probably biased.” Other
ber counts outside the 100–1300 ber/mm² range should be reported as having “greater
than optimal variability” and as being “probably biased.”
21. Calculate and report the concentration, (bers/cc), of bers in the air volume sampled, (L), using
the eective collection area of the lter, (approx. 385 mm² for a 25-mm lter):
NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
ASBESTOS and OTHER FIBERS by PCM: METHOD 7400, Issue 2, dated 15 August 1994 - Page 8 of 15
.
NOTE: Periodically check and adjust the value of , if necessary.
22. Report intralaboratory and interlaboratory relative standard deviations (from Step 11) with each set
of results.
NOTE: Precision depends on the total number of bers counted [1,16]. Relative standard deviation
is documented in references [1,15–17] for ber counts up to 100 bers in 100 graticule elds.
Comparability of interlaboratory results is discussed below. As a rst approximation, use
213% above and 49% below the count as the upper and lower condence limits for ber
counts greater than 20 (Figure 1).
EVALUATION OF METHOD:
Method Revisions:
This method is a revision of P&CAM 239 [10]. A summary of the revisions is as follows:
1. Sampling:
The change from a 37-mm to a 25-mm lter improves sensitivity for similar air volumes. The change
in ow rates allows for 2-m³ full-shift samples to be taken, providing that the lter is not overloaded
with non-brous particulates. The collection eciency of the sampler is not a function of ow rate in
the range 0.5 to 16 L/min [10].
2. Sample preparation technique:
The acetone vapor-triacetin preparation technique is a faster, more permanent mounting technique
than the dimethyl phthalate/diethyl oxalate method of P&CAM 239 [2,4,10]. The aluminum “hot
block” technique minimizes the amount of acetone needed to prepare each sample.
3. Measurement:
a. The Walton-Beckett graticule standardizes the area observed [14,18,19].
b. The HSE/NPL test slide standardizes microscope optics for sensitivity to ber diameter [4,14].
c. Because of past inaccuracies associated with low ber counts, the minimum recommended
loading has been increased to 100 bers/mm² lter area (a total of 78.5 bers counted in 100
elds, each with eld area = 0.00785 mm².) Lower levels generally result in an overestimate
of the ber count when compared to results in the recommended analytical range [20]. The
recommended loadings should yield intracounter
in the range of 0.10 to 0.17 [21–23].
Interlaboratory Comparability:
An international collaborative study involved 16 laboratories using prepared slides from the asbestos
cement, milling, mining, textile, and friction material industries [9]. The relative standard deviations (
)
varied with sample type and laboratory. The ranges were:
Rules Intralaboratory Interlaboratory Overall
AIA (NIOSH A Rules)* 0.12 to 0.40 0.27 to 0.85 0.46
Modied CRS (NIOSH B Rules)
†
0.11 to 0.29 0.20 to 0.35 0.25
*Under AIA rules, only bers having a diameter less than 3 µm are counted and bers attached to particles
larger than 3 µm are not counted. NIOSH A Rules are otherwise similar to the AIA rules.
†
See Appendix C.
A NIOSH study conducted using eld samples of asbestos gave intralaboratory in the range 0.17 to
0.25 and an interlaboratory of 0.45 [21]. This agrees well with other recent studies [9,14,16].
NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
ASBESTOS and OTHER FIBERS by PCM: METHOD 7400, Issue 2, dated 15 August 1994 - Page 9 of 15
At this time, there is no independent means for assessing the overall accuracy of this method. One
measure of reliability is to estimate how well the count for a single sample agrees with the mean count
from a large number of laboratories. The following discussion indicates how this estimation can be
carried out based on measurements of the interlaboratory variability, as well as showing how the results
of this method relate to the theoretically attainable counting precision and to measured intra- and
interlaboratory
. (NOTE: The following discussion does not include bias estimates and should not be
taken to indicate that lightly loaded samples are as accurate as properly loaded ones).
Theoretically, the process of counting randomly (Poisson) distributed bers on a lter surface will give
an
that depends on the number, , of bers counted:
.
Thus is 0.1 for 100 bers and 0.32 for 10 bers counted. The actual found in a number of studies is
greater than these theoretical numbers [17,19–21].
An additional component of variability comes primarily from subjective interlaboratory dierences. In
a study of ten counters in a continuing sample exchange program, Ogden [15] found this subjective
component of intralaboratory
to be approximately 0.2 and estimated the overall by the term:
.
Ogden found that the 90% condence interval of the individual intralaboratory counts in relation to
the means were +2
and −1.5 . In this program, one sample out of ten was a quality control sample.
For laboratories not engaged in an intensive quality assurance program, the subjective component of
variability can be higher.
In a study of eld sample results in 46 laboratories, the Asbestos Information Association also found
that the variability had both a constant component and one that depended on the ber count [14].
These results gave a subjective interlaboratory component of
(on the same basis as Ogden’s) for eld
samples of ca. 0.45. A similar value was obtained for 12 laboratories analyzing a set of 24 eld samples
[21]. This value falls slightly above the range of
(0.25 to 0.42 for 1984–85) found for 80 reference
laboratories in the NIOSH PAT program for laboratory-generated samples [17].
A number of factors inuence
for a given laboratory, such as that laboratory’s actual counting
performance and the type of samples being analyzed. In the absence of other information, such as
from an interlaboratory quality assurance program using eld samples, the value for the subjective
component of variability is chosen as 0.45. It is hoped that the laboratories will carry out the
recommended interlaboratory quality assurance programs to improve their performance and thus
reduce the
.
The above relative standard deviations apply when the population mean has been determined. It is
more useful, however, for laboratories to estimate the 90% condence interval on the mean count from
a single sample ber count (Figure 1). These curves assume similar shapes of the count distribution for
interlaboratory and intralaboratory results [16].
For example, if a sample yields a count of 24 bers, Figure 1 indicates that the mean interlaboratory
count will fall within the range of 227% above and 52% below that value 90% of the time. We can
apply these percentages directly to the air concentrations as well. If, for instance, this sample (24 bers
counted) represented a 500-L volume, then the measured concentration is 0.02 bers/mL (assuming
100 elds counted, 25-mm lter, 0.00785 mm² counting eld area). If this same sample were counted by
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ASBESTOS and OTHER FIBERS by PCM: METHOD 7400, Issue 2, dated 15 August 1994 - Page 10 of 15
a group of laboratories, there is a 90% probability that the mean would fall between 0.01 and 0.08 ber/
mL. These limits should be reported in any comparison of results between laboratories.
Note that the
of 0.45 used to derive Figure 1 is used as an estimate for a random group of laboratories.
If several laboratories belonging to a quality assurance group can show that their interlaboratory is
smaller, then it is more correct to use that smaller . However, the estimated of 0.45 is to be used in
the absence of such information. Note also that it has been found that can be higher for certain types
of samples, such as asbestos cement [9].
Quite often the estimated airborne concentration from an asbestos analysis is used to compare to a
regulatory standard. For instance, if one is trying to show compliance with an 0.5 ber/mL standard
using a single sample on which 100 bers have been counted, then Figure 1 indicates that the 0.5
ber/mL standard must be 213% higher than the measured air concentration. This indicates that if one
measures a ber concentration of 0.16 ber/mL (100 bers counted), then the mean ber count by a
group of laboratories (of which the compliance laboratory might be one) has a 95% chance of being
less than 0.5 bers/mL; i.e., 0.16 + 2.13 × 0.16 = 0.5.
It can be seen from Figure 1 that the Poisson component of the variability is not very important unless
the number of bers counted is small. Therefore, a further approximation is to simply use +213% and
−49% as the upper and lower condence values of the mean for a 100-ber count.
Figure 1. Interlaboratory precision of ber counts.
Number of fibers countedinasingle sample
95% probability mean countis above this level
Percent relative to single sample count
[subjectivecomponent (0.45)+Poisson component]
90% ConfidenceIntervalonMean Count
95% probability mean countis below this level
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The curves in Figure 1 are dened by the following equations:
and
,
where = subjective interlaboratory relative standard deviation, which is close to the total
interlaboratory when approximately 100 bers are counted,
= total bers counted on sample,
= lower 95% condence limit, and
= upper 95% condence limit.
Note that the range between these two limits represents 90% of the total range.
REFERENCES:
[1] Leidel, N. A., S. G. Bayer, R. D. Zumwalde, and K. A. Busch. USPHS/NIOSH Membrane Filter Method
for Evaluating Airborne Asbestos Fibers, U.S. Department of Health, Education, and Welfare, Publ.
(NIOSH) 79-127 (1979).
[2] Baron, P. A. and G. C. Pickford. “An Asbestos Sample Filter Clearing Procedure,” Appl. Ind. Hyg., 1,
169–171, 199 (1986).
[3] Occupational Safety and Health Administration, U.S. Department of Labor, Occupational Exposure
to Asbestos, Tremolite, Anthophyllite, and Actinolite Asbestos; Final Rules, 29 CFR Part 1910.1001
Amended June 20, 1986.
[4] Rooker, S. J., N. P. Vaughn, and J. M. LeGuen. “On the Visibility of Fibers by Phase Contrast
Microscopy,” Amer. Ind. Hyg. Assoc. J., 43, 505–515 (1982).
[5] Baron, P. and G. Deye, “Electrostatic Eects in Asbestos Sampling,” Parts I and II, Amer. Ind. Hyg. Assoc.
J., 51, 51–69 (1990).
[6] Johnston, A. M., A. D. Jones, and J. H. Vincent. “The Inuence of External Aerodynamic Factors
on the Measurement of the Airborne Concentration of Asbestos Fibers by the Membrane Filter
Method,” Ann. Occup. Hyg., 25, 309–316 (1982).
[7] Beckett, S.T., “The Eects of Sampling Practice on the Measured Concentration of Airborne
Asbestos,” Ann. Occup. Hyg., 21, 259–272 (1980).
[8] Jankovic, J. T., W. Jones, and J. Clere. “Field Techniques for Clearing Cellulose Ester Filters Used in
Asbestos Sampling,” Appl. Ind. Hyg., 1, 145–147 (1986).
[9] Crawford, N. P., H. L. Thorpe, and W. Alexander. “A Comparison of the Eects of Dierent Counting
Rules and Aspect Ratios on the Level and Reproducibility of Asbestos Fiber Counts,” Part I: Eects
on Level (Report No. TM/82/23), Part II: Eects on Reproducibility (Report No. TM/82/24), Institute of
Occupational Medicine, Edinburgh, Scotland (December, 1982).
[10] NIOSH Manual of Analytical Methods, 2nd ed., Vol. 1., P&CAM 239, U.S. Department of Health,
Education, and Welfare, Publ. (NIOSH) 77-157-A (1977).
[11] Revised Recommended Asbestos Standard, U.S. Department of Health, Education, and Welfare,
Publ. (NIOSH) 77-169 (1976); as amended in NIOSH statement at OSHA Public Hearing, June 21,
1984.
[12] Asbestos International Association, AIA Health and Safety Recommended Technical Method #1
(RTMI). “Airborne Asbestos Fiber Concentrations at Workplaces by Light Microscopy” (Membrane
Filter Method), London (1979).
[13] Abell, M., S. Shulman and P. Baron. “The Quality of Fiber Count Data,” Appl. Ind. Hyg., 4, 273–285
(1989).
[14] “A Study of the Empirical Precision of Airborne Asbestos Concentration Measurements in the
Workplace by the Membrane Filter Method,” Asbestos Information Association, Air Monitoring
Committee Report, Arlington, VA (June, 1983).
NIOSH Manual of Analytical Methods (NMAM), Fourth Edition
ASBESTOS and OTHER FIBERS by PCM: METHOD 7400, Issue 2, dated 15 August 1994 - Page 12 of 15
[15] McCrone, W., L. McCrone and J. Delly, “Polarized Light Microscopy,” Ann Arbor Science (1978).
[16] Ogden, T. L. “The Reproducibility of Fiber Counts,” Health and Safety Executive Research Paper 18
(1982).
[17] Schlecht, P. C. and S. A. Schulman. “Performance of Asbestos Fiber Counting Laboratories in the
NIOSH Prociency Analytical Testing (PAT) Program,” Am. Ind. Hyg. Assoc. J., 47, 259–266 (1986).
[18] Chateld, E. J. Measurement of Asbestos Fiber Concentrations in Workplace Atmospheres, Royal
Commission on Matters of Health and Safety Arising from the Use of Asbestos in Ontario, Study No.
9, 180 Dundas Street West, 22nd Floor, Toronto, Ontario, CANADA M5G 1Z8.
[19] Walton, W. H. “The Nature, Hazards, and Assessment of Occupational Exposure to Airborne Asbestos
Dust: A Review,” Ann. Occup. Hyg., 25, 115–247 (1982).
[20] Cherrie, J., A.D. Jones, and A.M. Johnston. “The Inuence of Fiber Density on the Assessment of
Fiber Concentration Using the membrane lter Method.” Am. Ind. Hyg. Assoc. J., 47(8), 465–74 (1986).
[21] Baron, P. A. and S. Shulman. “Evaluation of the Magiscan Image Analyzer for Asbestos Fiber
Counting.” Am. Ind. Hyg. Assoc. J., (in press).
[22] Taylor, D. G., P. A. Baron, S. A. Shulman and J. W. Carter. “Identication and Counting of Asbestos
Fibers,” Am. Ind. Hyg. Assoc. J. 45(2), 84–88 (1984).
[23] “Potential Health Hazards of Video Display Terminals,” NIOSH Research Report, June 1981.
[24] “Reference Methods for Measuring Airborne Man-Made Mineral Fibers (MMMF),” WHO/EURO
Technical Committee for Monitoring an Evaluating Airborne MMMF, World Health Organization,
Copenhagen (1985).
[25] Criteria for a Recommended Standard…Occupational Exposure to Fibrous Glass, U.S. Department
of Health, Education, and Welfare, Publ. (NIOSH) 77-152 (1977).
METHOD WRITTEN BY:
Paul A. Baron, Ph.D., NIOSH/DPSE.
APPENDIX A. CALIBRATION OF THE WALTON-BECKETT GRATICULE
Before ordering the Walton-Beckett graticule, the following calibration must be done to obtain a
counting area (
) 100 µm in diameter at the image plane. The diameter, (mm), of the circular counting
area and the disc diameter must be specied when ordering the graticule.
1. Insert any available graticule into the eyepiece and focus so that the graticule lines are sharp and
clear.
2. Set the appropriate interpupillary distance and, if applicable, reset the binocular head adjustment so
that the magnication remains constant.
3. Install the 40 to 45× phase objective.
4. Place a stage micrometer on the microscope object stage and focus the microscope on the
graduated lines.
5. Measure the magnied grid length of the graticule, (µm), using the stage micrometer.
6. Remove the graticule from the microscope and measure its actual grid length, (mm). This can best
be accomplished by using a stage tted with verniers.
7. Calculate the circle diameter, (mm), for the Walton-Beckett graticule:
.
Example: If = 112 µm, = 4.5 mm, and = 100 µm, then = 4.02 mm.
8. Check the eld diameter, (acceptable range 100 µm ± 2 µm) with a stage micrometer upon receipt
of the graticule from the manufacturer. Determine eld area (acceptable range 0.00754 mm² to
0.00817 mm²).
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APPENDIX B. COMPARISON OF COUNTING RULES
Figure 2 shows a Walton-Beckett graticule as seen through the microscope. The rules will be discussed
as they apply to the labeled objects in the gure.
Figure 2. Walton-Beckett graticule with bers.
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These rules are sometimes referred to as the “A” rules:
Object Count Discussion
1 1 ber Optically observable asbestos bers are actually bundles of ne brils. If the
brils seem to be from the same bundle, the object is counted as a single
ber. Note, however, that all objects meeting length and aspect ratio criteria
are counted whether or not they appear to be asbestos.
2 2 bers If bers meeting the length and aspect ratio criteria (length >5 µm and
length-to-width ratio > 3 to 1) overlap, but do not seem to be part of the
same bundle, they are counted as separate bers.
3 1 ber Although the object has a relatively large diameter (>3 µm), it is counted as -
ber under the rules. There is no upper limit on the ber diameter in the count-
ing rules. Note that ber width is measured at the widest compact section of
the object.
4 1 ber Although long ne brils may extend from the body of a ber, these brils are
considered part of the ber if they seem to have originally been part of the
bundle.
5 Do not count If the object is ≤ 5 µm long, it is not counted.
6 1 ber A ber partially obscured by a particle is counted as one ber. If the ber ends
emanating from a particle do not seem to be from the same ber and each
end meets the length and aspect ratio criteria, they are counted as separate
bers.
7 ½ ber A ber which crosses into the graticule area one time is counted as ½ ber.
8 Do not count Ignore bers that cross the graticulate boundary more than once.
9 Do not count Ignore bers that lie outside the graticule boundary.
APPENDIX C. ALTERNATE COUNTING RULES FOR NON-ASBESTOS FIBERS
Other counting rules may be more appropriate for measurement of specic non-asbestos ber types,
such as brous glass. These include the “B” rules given below (from NIOSH Method 7400, Revision #2,
dated 8/15/87), the World Health Organization reference method for man-made mineral ber [24], and
the NIOSH brous glass criteria document method [25]. The upper diameter limit in these methods
prevents measurements of non-thoracic bers. It is important to note that the aspect ratio limits
included in these methods vary. NIOSH recommends the use of the 3:1 aspect ratio in counting bers.
It is emphasized that hybridization of dierent sets of counting rules is not permitted. Report
specically which set of counting rules are used with the analytical results.
“B” Counting Rules
1. Count only ends of bers. Each ber must be longer than 5 µm and less than 3 µm diameter.
2. Count only ends of bers with a length-to-width ratio equal to or greater than 5:1.
3. Count each ber end which falls within the graticule area as one end, provided that the ber meets
rules 1 and 2 above. Add split ends to the count as appropriate if the split ber segment also meets
the criteria of rules 1 and 2 above.
4. Count visibly free ends which meet rules 1 and 2 above when the ber appears to be attached to
another particle, regardless of the size of the other particle. Count the end of a ber obscured by
another particle if the particle covering the ber end is less than 3 µm in diameter.
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5. Count free ends of bers emanating from large clumps and bundles up to a maximum of 10 ends (5
bers), provided that each segment meets rules 1 and 2 above.
6. Count enough graticule elds to yield 200 ends. Count a minimum of 20 graticule elds. Stop at 100
graticule elds, regardless of count.
7. Divide total end count by 2 to yield ber count.
APPENDIX D. EQUIVALENT LIMITS OF DETECTION AND QUANTITATION
Fiber density on lter* Fiber concentration in air, f/cc
Fibers per 100 elds Fibers/mm² 400-L air sample 1000-L air sample
200 255 0.25 0.10
100 127 0.125 0.05
LOQ 80.0 102 0.10 0.04
50 64 0.0625 0.025
25 32 0.03 0.0125
20 25 0.025 0.010
10 12.7 0.0125 0.005
8 10.2 0.010 0.004
LOD 5.5 7 0.00675 0.0027
*Assumes 385 mm² eective lter collection area, and eld area = 0.00785 mm², for relatively “clean” (little
particulate aside from bers) lters.
