Interlaboratory evaluation of endotoxin analyses in agricultural dusts—comparison of LAL assay and mass spectrometry

Stephen J. Reynolds; Donald K. Milton; Dick Heederik; Peter S. Thorne; Kelley J. Donham; Elizabeth A. Croteau; Kevin M. Kelly; Jeroen Douwes; Daniel Lewis; Mike Whitmer; Ian Connaughton; Sharon Koch; Per Malmberg; Britt-Marie Larsson; Jim Deddens; Anita Saraf; Lennart Larsson

doi:10.1039/B509256F

Interlaboratory evaluation of endotoxin analyses in agricultural dusts—comparison of LAL assay and mass spectrometry

Stephen J. Reynolds,*^a Donald K. Milton,^b Dick Heederik,^c Peter S. Thorne,^d Kelley J. Donham,^e Elizabeth A. Croteau,^d Kevin M. Kelly,^d Jeroen Douwes,^c Daniel Lewis,^e Mike Whitmer,^e Ian Connaughton,^f Sharon Koch,^f Per Malmberg,^g Britt-Marie Larsson,^g Jim Deddens,^h Anita Sarafⁱ and Lennart Larssonⁱ

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* Corresponding authors

^a Department of Environmental and Radiological Health Sciences, Colorado State University, Ft. Collins, USA
E-mail: Stephen.Reynolds@Colostate.edu
Fax: +1 970-491-2940

^b Department of Environmental Health, Harvard School of Public Health, 665 Huntington Ave, Boston, USA

^c Institute for Risk Assessment Sciences (IRAS), Division of Environmental and Occupational Health, University of Utrecht, PO Box 80176, Utrecht, The Netherlands

^d College of Public Health, Department of Occupational and Environmental Health, The University of Iowa, Iowa City, USA

^e Division of Respiratory Disease Studies/NIOSH, 1095 Willowdale Road, Morgantown, USA

^f Bendigo Agricultural Centre, Bendigo Mail Centre, Victoria, Australia

^g National Institute for Working Life, Stockholm, Sweden

^h Department of Mathematical Sciences, University of Cincinnati, Cincinnati, USA

ⁱ Department of Laboratory Medicine, Section of Medical Microbiology, University of Lund, Sölvegatan 23, Lund, Sweden

Abstract

Endotoxin exposure is associated with wheeze and asthma morbidity, while early life exposure may reduce risk of allergy and asthma. Unfortunately, it is difficult to compare endotoxin results from different laboratories and environments. We undertook this study to determine if lipopolysaccharide (LPS) extraction efficiency could account for differences among laboratories. We generated and collected aerosols from chicken and swine barns, and corn processing. We randomly allocated side-by-side filter samples to five laboratories for Limulus assay of endotoxin. Lyophilized aliquots of filter extracts were analyzed for 3-hydroxy fatty acids (3-OHFAs) as a marker of LPS using gas chromatography-mass spectrometry. There were significant differences in endotoxin assay and GC-MS (LPS) results between laboratories for all dust types (p < 0.01). Patterns of differences between labs varied by dust type. Relationships between assay and GC/MS results also depended on dust type. The percentages of individual 3-OHFA chain lengths varied across labs (p < 0.0001) suggesting that each lab recovered a different fraction of the LPS available. The presence of large amounts of particle associated LPS and absence of a freezing thawing cycle were associated with lower correlations between LPS and bioactivity, consistent with an absence of Limulus response to cell-bound endotoxin. These data suggest that extraction methods affect endotoxin measurements. The LAL methods may be most suitable when comparing exposures within similar environments; GC-MS offers additional information helpful in optimizing sample treatment and extraction. GC-MS may be of use when comparing across heterogeneous environments and should be considered for inclusion in future studies of human health outcomes.

Journal of Environmental Monitoring

Interlaboratory evaluation of endotoxin analyses in agricultural dusts—comparison of LAL assay and mass spectrometry

Abstract

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Interlaboratory evaluation of endotoxin analyses in agricultural dusts—comparison of LAL assay and mass spectrometry

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