A new method for simultaneous immunodetection and morphologic identification of individual sources of pollen allergens

Background: Exposure to outdoor allergens has commonly been estimated by co llecting airborne particles with a Hirst-type spore trap and then using mor phologic criteria to identify the intact pollen grains and fungal spores th at are recognized as allergen sources. Several antibody-based blotting or f ixation methods have also been developed that enable the counting of amorph ous airborne particles carrying allergen, but none of these methods allow t he ready association of the released allergen with the morphologically iden tifiable particle of origin. A method has been developed that uses pressure -sensitive adhesive tape to sample the airborne particles and then allows t he immunoidentification of the specific particles that are the allergen sou rces. Objective: Our purpose was to visualize and immunostain the particles carry ing pollen allergen that are collected with a volumetric spore trap. Methods: A Burkard sampler was used to collect airborne particles onto pres sure-sensitive adhesive tapes. The particles were permanently fixed between the tape and a protein-binding membrane when the tape was laminated with t he membrane. Allergens that elute from the particles onto the membrane were detected with a range of antibodies. Both the particle and associated immu nostained allergen were viewed through the transparent tape for final micro scopic identification. Results: Polyclonal and monoclonal antibodies and IgE from allergic patient s stained allergens in the periphery of particles collected on the tapes. I ndividual pollen grains and paucimicronic particles were seen with halos of immunostained allergen surrounding them. When IgE was used, the density of immunostaining in the halo surrounding Lolium perenne pollen grains was fo und to be proportional to the level of Lolium-specific IgE. The method is h ighly sensitive and can be used to detect different airborne particles that carry allergen. Both the particle and the immunostaining can be subjected to a range of simple measurement techniques. Conclusion: Individual particles carrying allergens and antigens were visua lized. These particles included intact pollen grains, paucimicronic particl es, and fungal spores.