Comparison of the limulus amebocyte lysate (LAL) and recombinant factor C (rFC) endotoxin bioassays: characterization of aerosols and settled dust on a sheep farm

Abstract

The goals of this study are: 1) To optimize an extraction protocol for bulk sheep dust analysis by rFC; 2) To compare the traditional chromogenic LAL to the newer fluorometric rFC assay; 3) To compare the effects of the two most widely used extraction media, pyrogen-free water (PFW) and pyrogen-free water with 0.05% Tween 20 (PFW-Tween 20), on the assay outcomes; and 4) To characterize endotoxin exposure in a sheep farm environment. Settled and airborne dust samples were collected from a sheep farm for analysis. Settled dust was analyzed with endpoint fluorogenic rFC and kinetic chromogenic LAL in two different laboratories. A total of 34 stationary airborne dust samples (32 matched) were collected with Institute of Occupational Medicine (IOM) inhalable dust samplers, using polyvinyl chloride (PVC) filters. Airborne samples were analyzed with endpoint fluorogenic rFC and endpoint chromogenic LAL in the same laboratory. The results of the bulk dust analysis showed a significant difference in polystyrene (PS) tubes and polypropylene (PP) tubes used for extraction indicating that PS tubes yielded higher endotoxin levels (t = 3.82, p = 0.000). No difference in endotoxin levels was found for centrifugation and spike recovery was closest to 100% for extraction with PFW. The results of the airborne dust study showed a strong positive correlation between the rFC and LAL assays with PFW-Tween 20 extraction. All samples extracted in PFW-Tween 20 had a higher endotoxin recovery compared with those extracted in PFW for both the rFC and LAL assays.