Development of a Sandwich ELISA to Measure Exposure to Occupational Cow Hair Allergens

Abstract

Background: Cow hair and dander are important inducers of occupational allergies in cattle-exposed farmers. To estimate allergen exposure in farming environments, a sensitive enzyme immunoassay was developed to measure cow hair allergens. Methods: A sandwich ELISA was developed using polyclonal rabbitantibodies against a mixture of hair extracts from different cattle breeds. To assess the specificity of the assay, extracts from other mammalian epithelia, mites, molds and grains were tested. To validate the new assay, cow hair allergens were measured in passive airborne dust samples from the stables and homes of farmers. Dust was collected with electrostatic dust fall collectors (EDCs). Results: The sandwich ELISA was found to be very sensitive (detection limit: 0.1 ng/ml) and highly reproducible, demonstrating intra- and interassay coefficients of variation of 4 and 10%, respectively. The assay showed no reactivity with mites, molds and grains, but some cross-reactivity with other mammalian epithelia, with the strongest reaction with goat. Using EDCs for dust sampling, high concentrations of bovine allergens were measured in cow stables (4,760–559,400 µg/m²). In addition, bovine allergens were detected in all areas of cattle farmer dwellings. A large variation was found between individual samples (0.3–900 µg/m²) and significantly higher values were discovered in changing rooms. Conclusion: The ELISA developed for the detection of cow hair proteins is a useful tool for allergen quantification in occupational and home environments. Based on its low detection limit, this test is sensitive enough to detect allergens in passive airborne dust.

Allergen Levels in the Hair of Different Cattle Breeds

“…In stables, the levels of bovine hair allergens in airborne dust can reach immensely high values. In our previous report, the median allergen concentration was 14 mg/g dust, and allergen levels differed by about 100-fold between cow stables (all with Simmental cattle), ranging from 5,000 to 560,000 μg/m 2 [11] . It is likely that some proportion of the airborne allergen produced by livestock settles back onto the fur during the sedimentation process.…”

“…Bovine hair allergen concentrations were determined via a sandwich ELISA as described by Zahradnik et al [11] . The assay was developed using affinity-purified polyclonal antibodies against the total soluble antigen spectrum extracted from bovine hair.…”

“…The assay was developed using affinity-purified polyclonal antibodies against the total soluble antigen spectrum extracted from bovine hair. The bovine hair extract was previously examined for the presence of allergens using an IgE immunoblot with sera from several cow dander-sensitized patients, which recognized almost the entire protein pattern [11] .…”

“…SDS-PAGE analysis of cow hair extracts has shown nearly identical protein patterns with a dominant band at 20 kDa without any striking breed-or gender-specific differences. Some variations have appeared in the intensities of single bands of protein profiles [11] . Using immunoelectrophoresis methods, only minor differences in allergen composition have been identified upon analysis of material from 4 different cattle breeds [2] .…”

Exposure levels, determinants and IgE mediated sensitization to bovine allergens among Danish farmers and non-farmers

“…Few studies have quantitatively assessed bovine allergens in the environment, and they all found high levels of allergens in cow stables (Berger et al, 2005;Virtanen et al, 1988;Zahradnik et al, 2011). Zahradnik et al (2011) reported median levels of bovine hair allergen in cow stables to be 51,700 g/m 2 .…”