The windborne pollen grains of many trees and grasses contain a number of highly water-soluble (glyco)proteins which upon moistening, e.g. on the human respiratory mucosa, rapidly diffuse out of the pollen grain. In susceptible individuals, they may cause allergic reactions. Because of their rapid release from the pollen, pollen allergens were expected to be located in the outermost layers of the pollen grain, i.e. on the surface and in the wall (exine, intine). First attempts to localize allergens by microscopic methods in the pollen grain supported this view. However, since conventional preparation methods were used, artificial mobilization of allergens could not be excluded. Based on new, completely anhydrous preparation protocols and on the immunogold-labeling technique, pollen allergens were electron-microscopically shown to be located in the interior of the pollen grain, i.e. in the cytoplasm, often within ribosome-rich areas. This unexpected result was obtained in all strictly anhydrously prepared pollen species. The biological function of these allergens is largely unknown, however, clues as to possible cellular functions have been obtained for the birch pollen major allergen Bet v 1.