Localization of Four Allergens in Artemisia Pollen by Immunofluorescent Antibodies

Abstract

Background: Artemisia pollens have a high potential to induce allergic symptoms. Seven allergen components have been identified, but only Art v 7 has been localized in the pollen grain. This study aimed to localize the allergens in the pollen grains of 4 Artemisia spp. Methods: Pollen extracts from 2 Chinese Artemisia spp., A. argyi and A. annua, were used to immunize BALB/c mice. Recombinant Art v 1 and Art v 3 allergens were used to select specific monoclonal antibodies (mAbs). Three mAbs were used to purify the natural allergens and were then analyzed by mass spectrometry. As reported previously, polyclonal antibodies were obtained from rabbits immunized with 3 synthesized peptides of Art an 7. Using conventional histology procedures with pollens from 4 Artemisia spp. (A. argyi, A. annua, A. capilaris, and A. sieversiana), allergen images were observed and recorded by fluorescence and confocal laser microscopy. Results: We obtained 2 specific mAbs against Art v 1, 1 against Art v 2, and 4 against Art v 3 homologs. The Art v 1 and Art v 3 homologs were mainly located on the pollen walls, and the Art v 7 homologous protein was localized intracellularly around nuclei. The location of the Art v 2 homologous protein varied across species, being intracellular around nuclei for A. annua and A. argyi, and in both the pollen wall and around nuclei for A. capilaris and A. sieversiana. Conclusions: Four mugwort allergens were localized in the pollen, and the major Art v 1 and Art v 3 allergens were located mainly in the pollen wall.

Variation in IgE binding potencies of seven Artemisia species depending on content of major allergens

“…Monoclonal antibodies (mAb) used in this study were from previous research [27 ]. A7‐G4‐E6 specific to Art v 1, C9‐C1 to Art v 2, and A2‐B8 to Art v 3, were used to purify three groups of allergens from six selected Artemisia spp.…”

“…A7‐G4‐E6 specific to Art v 1, C9‐C1 to Art v 2, and A2‐B8 to Art v 3, were used to purify three groups of allergens from six selected Artemisia spp. from China as described previously [27 ]. LC–MS/MS (Thermo Scientific Q Exactive) was used for identity‐matching of the purified proteins to deduced allergens from each species.…”

Allergenic Properties and Molecular Characteristics of PR-1 Proteins

“…Artemisia vulgaris ( 27 ) belongs to one of the largest families of flowering plants, the Compositae or Asteraceae . It is a weed widely spread in temperate regions and subtropics of Europe, North America, and Asia ( 28 , 29 ). Among these, Art v 1, Art v 2, Art v 3, and Art v 7 have been shown to be the most relevant allergens ( 28 , 29 ).…”

“…It is a weed widely spread in temperate regions and subtropics of Europe, North America, and Asia ( 28 , 29 ). Among these, Art v 1, Art v 2, Art v 3, and Art v 7 have been shown to be the most relevant allergens ( 28 , 29 ). However, the function of PR-1 homologous Art v 2 remains unknown ( 14 ).…”

“…Artemisia pollens have been recognized as a major cause for seasonal allergic respiratory disease worldwide, especially asthma ( 28 , 34 ). It is relevant along with the Asia–Europe silk-road and in northwestern European countries.…”