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Background: Vine culture is widely distributed in La Rioja (37% of the crops), which involves exposure of the general population to this plant’s pollen. The aim of this study was to investigate the prevalence of sensitization to Vitis vinifera pollen in the general population with respiratory allergy and to identify the allergens involved. Materials and methods: Patients who came to the hospital between September 2019 and January 2020, with suspected respiratory allergy were included in the study. All of them were skin prick tested with a panel of standardized aeroallergens, profilin, LTP, V. vinifera pollen extract and prick-prick with fresh grapes. In vitro study included specific IgE by ImmunoCap and ELISA, allergenic profile by immunoblot with individual sera from patients positive to V. vinifera pollen extract and immunoblot 2D with a pool of sera. The spots recognized by IgE were identified by mass spectrometry. Results: A total of 151 patients were included. Of them, 124 were positive to some of the allergens tested. Thirty-four (27.4%) were positive to vine pollen by skin prick test. After the serological study, 20 had positive results. Five vine pollen allergens were identified, and profilin was the most prevalent (30%). The other four allergens could be considered specific to this pollen. Conclusion: A high sensitization to vine pollen was detected in the general population in a vineyard area. The clinical relevance is unknown due to the sensitization to other pollens in the vine pollen positive patients. Five new vine pollen allergens were identified.