Development of a comprehensive protein microarray for immunoglobulin E profiling in horses with severe asthma

Moore?Colyer, Meriel; White, Samuel; Moore-Colyer, Meriel; Marti, Eliane; Co�etil, Laurent; Hannant, Duncan; Richard, Eric A.; Alcocer, Marcos

Abstract

Background
Severe asthma in horses, known as severe equine asthma (SEA), is a prevalent, performance‐limiting disease associated with increased allergen‐specific immunoglobulin E (IgE) against a range of environmental aeroallergens.

Objective
To develop a protein microarray platform to profile IgE against a range of proven and novel environmental proteins in SEA‐affected horses.

Animals
Six SEA‐affected and 6 clinically healthy Warmblood performance horses.

Methods
Developed a protein microarray (n = 384) using protein extracts and purified proteins from a large number of families including pollen, bacteria, fungi, and arthropods associated with the horses, environment. Conditions were optimized and assessed for printing, incubation, immunolabeling, biological fluid source, concentration techniques, reproducibility, and specificity.

Results
This method identified a number of novel allergens, while also identifying an association between SEA and pollen sensitization. Immunolabeling methods confirmed the accuracy of a commercially available mouse anti‐horse IgE 3H10 source (R2 = 0.91). Biological fluid source evaluation indicated that sera and bronchoalveolar lavage fluid (BALF) yielded the same specific IgE profile (average R2 = 0.75). Amicon centrifugal filters were found to be the most efficient technique for concentrating BALF for IgE analysis at 40‐fold. Overnight incubation maintained the same sensitization profile while increasing sensitivity. Reproducibility was demonstrated (R2 = 0.97), as was specificity using protein inhibition assays. Arthropods, fungi, and pollens showed the greatest discrimination for SEA.

Conclusions and Clinical Importance
We have established that protein microarrays can be used for large‐scale IgE mapping of allergens associated with the environment of horses. This technology provides a sound platform for specific diagnosis, management, and treatment of SEA.

Citation

Moore‐Colyer, M., White, S., Moore-Colyer, M., Marti, E., Coüetil, L., Hannant, D., Richard, E. A., & Alcocer, M. (2019). Development of a comprehensive protein microarray for immunoglobulin E profiling in horses with severe asthma. Journal of Veterinary Internal Medicine, 33(5), 2327-2335. https://doi.org/10.1111/jvim.15564

• Journal Article Type: Article
• Acceptance Date: Jul 3, 2019
• Online Publication Date: Aug 20, 2019
• Publication Date: Aug 20, 2019
• Deposit Date: Jul 8, 2019
• Publicly Available Date: Aug 28, 2019
• Journal: Journal of Veterinary Internal Medicine
• Print ISSN: 0891-6640
• Electronic ISSN: 1939-1676
• Publisher: Wiley Open Access
• Peer Reviewed: Peer Reviewed
• Volume: 33
• Issue: 5
• Pages: 2327-2335
• DOI: https://doi.org/10.1111/jvim.15564
• Public URL: https://nottingham-repository.worktribe.com/output/2284780
• Publisher URL: https://onlinelibrary.wiley.com/doi/full/10.1111/jvim.15564
• Contract Date: Jul 8, 2019