Timothy grass (Phleum pratense L.) pollen as allergen carriers and initiators of an allergic response

Contrary to indoor allergen exposure (e.g. house dust mite), there is no re liable quantitative association between pollen exposure and symptoms of all ergic diseases. Therefore we studied localization and release of major alle rgens from timothy grass (Phleum pratense L.) pollen using different method s and pollen grain sources. Localization of major allergens Phl p 5 and Phl p 1 was visualized by field emission scanning electron microscopy after an hydrous fixation and immunogold silver staining in a three-dimensional reco nstruction; Phl p 5 was found in the cytoplasm and on the exine, Phl p 1 in the intine. No allergens were found inside the starch granules. Allergen l iberation from pollen grains was studied in vitro under physiological condi tions (30 min, 37 degrees C) at pH 6.0, 7.4 and 9.0. Besides total protein measurements in the supernatant, major allergens were determined by immunob lot, Phl p 5 was quantitated by ELISA. There were striking differences in t otal protein and major allergen release between freshly collected and comme rcially available grass pollen grains as well as among freshly collected po llen between rural meadows and areas near high-traffic roads. There was a s ignificantly different release of total protein being lowest in supernatant s from commercially available pollen grains (rural/traffic vs. commercial, p<0.001), and of Phl p 5 major allergen (rural>traffic>commercial, p<0.005) . Therefore, allergen bioavailability seems to be an important parameter in order to establish reliable dose-response relationships for the outdoor al lergen response. Pollen grains incubated in aqueous protein-free buffer sol ution were also found to secrete significant amounts of eicosanoids namely prostaglandin E2 and leukotriene B4. Pollen grains thus do not act only as allergen carriers but also might have important implications on early event s as initiators of allergy.