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. 2025 Oct 14;20(1):35.
doi: 10.1186/s12995-025-00476-3.

Preparation methodology evaluation of rat pulmonary tissues containing mineral fibers following inhalation exposure to Libby amphibole asbestos

Jamie S Richey  1 , John R Shaw  1 , Amit Gupta  1 , Dawn M Fallacara  1 , Barney R Sparrow  1 , Anbo Wang  1 , Karen E Elsass  2 , Georgia K Roberts  3 , Pei-Li Yao  4 , Matthew D Stout  3 , Benjamin J Ellis  5 , Robyn L Ray  6
Affiliations

Preparation methodology evaluation of rat pulmonary tissues containing mineral fibers following inhalation exposure to Libby amphibole asbestos

Jamie S Richey et al. J Occup Med Toxicol. .

Abstract

Background: Inhaled mineral fibers including asbestos are associated with lung cancer and pleural disease. In this study, we evaluated methodologies for mineral fiber isolation with subsequent physical and chemical characterization from pulmonary tissues of rats exposed to Libby amphibole asbestos 2007 (LA 2007) fibers via repeated nose-only inhalation. At the completion of the exposures, lungs were collected either as is or instilled with liquid agarose to produce a pulmonary cast. To extract fibers, lung tissue with and without pulmonary casts were further processed by either high temperature ashing or chemical digestion. The use of liquid agarose to produce pulmonary casts was discontinued after the first study assessment as no fibers were present in the pleural cavity for evaluation. Fibers isolated from the lungs were analyzed by transmission electron microscopy (TEM) coupled with selected area electron diffraction (SAED) and energy dispersive X-ray spectroscopy (EDS) for physical and chemical characterization. The bulk LA 2007 test material was also analyzed to provide comparison of fiber dimensions and chemical composition of the fibers introduced during exposure. Particular interest was focused on the comparison between high temperature ashing and chemical digestion extraction methodologies.

Results: Chemical digestion of lung tissue with and without pulmonary casts resulted in fiber dimensions and chemical profiles similar to the bulk LA 2007 test chemical and exposure atmosphere. Conversely, high temperature ashing resulted in degraded fibers with chemically altered profiles.

Conclusions: Based on the findings in this study, chemical digestion of lung tissue is the preferred preparation method for the isolation of inhaled mineral fibers for lung burden analysis.

Keywords: Asbestos; Chemical digestion; Fiber burden; High temperature ashing; Inhaled mineral fiber; Libby amphibole.

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Conflict of interest statement

Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
TEM Images of Ashed Lung Fibers Showing NAM Fibers (A-D) and LA Fibers (E-F)
Fig. 2
Fig. 2
TEM Images of Fibers from Chemical Digested Lungs Classified as LA (A-D)
Fig. 3
Fig. 3
EDS Spectra (A) Pre-aerosolized LA, (B) Ashed Lung NAM Fibers, (C) Digested Lung LA Fibers
Fig. 4
Fig. 4
Length and Width Histograms: (A-B) Pre-Aerosolized LA 2007, (C-D) Ashed Lung NAM Fibers, (E-F) Digested Lung LA Fibers
See this image and copyright information in PMC

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