Evaluation of the dark-medium objective lens in counting asbestos fibers by phase-contrast microscopy

Our plain-English summary of Evaluation of the dark-medium objective lens in counting asbestos fibers by phase-contrast microscopy, by E.G. Lee, J.H. Nelson, M.L. Kashon, et al (The Annals of Occupational Hygiene, 2015).

The study

• Authors: E.G. Lee, J.H. Nelson, M.L. Kashon, M. Harper
• Journal: The Annals of Occupational Hygiene
• Published: 2015;59(5):616-28
• Cited by: 4 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

A Japanese round-robin study revealed that analysts who used a dark-medium (DM) objective lens reported higher fiber counts from American Industrial Hygiene Association (AIHA) Proficiency Analytical Testing (PAT) chrysotile samples than those using a standard objective lens, but the cause of this difference was not investigated at that time. The purpose of this study is to determine any major source of this difference by performing two sets of round-robin studies. For the first round-robin study, 15 AIHA PAT samples (five each of chrysotile and amosite generated by water-suspended method, and five chrysotile generated by aerosolization method) were prepared with relocatable cover slips and examined by nine laboratories. A second round-robin study was then performed with six chrysotile field sample slides by six out of nine laboratories who participated in the first round-robin study. In addition, two phase-shift test slides to check analysts' visibility and an eight-form diatom test plate to compare resolution between the two objectives were examined. For the AIHA PAT chrysotile reference slides, use of the DM objective resulted in consistently higher fiber counts (1.45 times for all data) than the standard objective (P-value < 0.05), regardless of the filter generation (water-suspension or aerosol) method. For the AIHA PAT amosite reference and chrysotile field sample slides, the fiber counts between the two objectives were not significantly different. No statistically significant differences were observed in the visibility of blocks of the test slides between the two objectives. Also, the DM and standard objectives showed no pattern of differences in viewing the fine lines and/or dots of each species images on the eight-form diatom test plate. Among various potential factors that might affect the analysts' performance of fiber counts, this study supports the greater contrast caused by the different phase plate absorptions as the main cause of high counts for the AIHA PAT chrysotile slides using the DM objective. The comparison of fiber count ratios (DM/standard) between the AIHA PAT chrysotile samples and chrysotile field samples indicates that there is a fraction of fibers in the PAT samples approaching the theoretical limit of visibility of the phase-contrast microscope with 3-degree phase-shift. These fibers become more clearly visible through the greater contrast from the phase plate absorption of the DM objective. However, as such fibers are not present in field samples, no difference in counts between the two objectives was observed in this study. The DM objective, therefore, could be allowed for routine fiber counting as it will maintain continuity with risk assessments based on earlier phase-contrast microscopy fiber counts from field samples. Published standard methods would need to be modified to allow a higher aperture specification for the objective.

Published by Oxford University Press on behalf of the British Occupational Hygiene Society 2015.

In plain English

This laboratory methods study looked at how the choice of microscope lens affects counts of asbestos fibers under phase-contrast microscopy, the standard method for measuring airborne asbestos. It did not study health effects. Nine laboratories counted fibers on 15 proficiency-testing reference slides (ten chrysotile, five amosite). All of them used the same circulated Nikon microscope, counting once with a standard objective lens (DLL, numerical aperture 0.75) and once with a 'dark-medium' (DM) lens (numerical aperture 0.95). Six of the labs then counted six chrysotile slides made from older workplace air samples whose site is unknown. With the DM lens, counts on the chrysotile reference slides were significantly higher; the authors give 1.45 times for all data. Counts did not differ significantly between the two lenses for the amosite reference slides or the chrysotile field slides. Test slides and a diatom test plate showed no consistent differences between the lenses in visibility or resolution. The authors concluded that the greater contrast of the DM lens was the main cause of the higher counts. They concluded the DM lens could be allowed for routine counting, although published standard methods would need to be changed. They noted that the field results came from only six slides from a single site.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Interlaboratory (round-robin) comparison of two microscope objective lenses, done in two rounds, with supporting test-slide, diatom test plate and transmission electron microscopy (TEM) checks
• Who was studied: Nine volunteer laboratories examined 15 AIHA Proficiency Analytical Testing reference slides: five chrysotile made by water suspension, five chrysotile made by aerosolization, and five amosite made by water suspension. Six of the nine laboratories then examined six chrysotile field sample slides. These were leftover samples collected many years earlier; the workplace location and type are unknown. Dates of the round-robins are not given.
• Exposure: The study compared counts of chrysotile and amosite fibers made with a dark-medium (DM) objective (Plan Apochromatic, x40, numerical aperture 0.95) and a standard DLL objective (Plan Fluorite, x40, numerical aperture 0.75) on a Nikon 55i phase-contrast microscope. Counting followed the NIOSH 7400 'A' rules plus fiber width under 3 micrometers.
• Main results:
  • AIHA PAT chrysotile reference slides: the DM objective gave consistently higher fiber counts than the standard objective, 1.45 times for all data (P less than 0.05), regardless of how the filters were generated.
  • Water-suspension chrysotile reference slides (9 labs): median ratio of DM to DLL counts 1.30 (range 0.75 to 3.29). P = 0.0066; model estimates 70 fibers with DM versus 58 with DLL.
  • Aerosol-generated chrysotile reference slides (9 labs): median ratio of DM to DLL counts 1.37 (range 0.63 to 4.08). P = 0.0001; model estimates 81 fibers with DM versus 63 with DLL.
  • Amosite reference slides (9 labs) and chrysotile field slides (6 labs): fiber counts were not significantly different between the two objectives. For amosite, the median ratio was 1.00, P = 0.9778, estimates 112 versus 112. For chrysotile field slides, the median ratio was 1.03, P = 0.5058, estimates 71 versus 68.
  • On the chrysotile reference slides, counts with either lens were below the 'verified fiber' counts set by two experienced analysts. Median count divided by verified fibers: water-suspension slides 0.62 with DM and 0.47 with DLL; aerosol slides 0.64 with DM and 0.53 with DLL.
  • Phase-shift test slides: visibility of the test slide blocks did not differ significantly between the DM and DLL lenses, or between x10 and x12.5 eyepieces.
  • Eight-form diatom test plate (8 analysts): responses on whether the DM or DLL lens resolved fine lines and dots better showed no pattern.
  • TEM of the chrysotile reference filters: average fiber width was 0.181 micrometers for water-suspension samples (827 fibers examined) and 0.182 micrometers for aerosol samples (635 fibers examined).
• The authors' conclusion: The authors concluded that the higher contrast from the DM objective's greater phase plate absorption is the main factor behind the higher chrysotile reference slide counts. They concluded the DM objective could be allowed for routine fiber counting while keeping continuity with risk assessments based on earlier phase-contrast counts of field samples. They added that published standard methods would need to be modified to allow a higher aperture.
• Limitations the authors state:
  • The field sample finding rests on only six chrysotile slides from a single, unknown field site, all with fiber densities below 230 fibers per square millimeter.
  • Fiber counts with the two lenses may still differ slightly depending on how tired the analysts' eyes were.
  • One lab (Lab F) did not finish counting the chrysotile reference slides. Another lab (Lab I) was excluded from the test slide analysis because it misunderstood the guidance.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Amosite, Chrysotile.

Medical terms in this study, explained in our Lexicon: Industrial Hygiene, Chrysotile, Amosite, P-Value, Phase Plate, Fiber Counting.

Related studies

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• A technical comparison of evaluating asbestos concentration by phase-contrast microscopy (PCM), scanning electron microscopy (SEM), and analytical transmission electron microscopy (ATEM) as illustrated from data generated from a case report — R.F. Dodson, S.P. Hammar, et al. (2008)
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• Rapid fiber-detection technique by artificial intelligence in phase-contrast microscope images of simulated atmospheric samples — T. Yamamoto, K. Iwasaki, et al. (2024)
• Automated counting of airborne asbestos fibers by a high-throughput microscopy (HTM) method — M.O. Cho, S. Yoon, et al. (2011)

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