Carrot (Daucus carota) allergy is caused by the major carrot allergen Dau c 1, which is a mixture of several isoallergens and variants with sequence identities of >67% or >90%, respectively. However, little is known about the qualitative and quantitative composition of natural Dau c 1.Mass spectrometry of isolated natural Dau c 1 reveals the existence of several yet unknown Dau c 1-like proteins. The study expresses four Dau c 1-like proteins in Escherichia coli. Two of the purified proteins, designated Dau c 1.0501 and 1.0601, exhibit sequence identities to Dau c 1.0101 and 1.0401 between 54% and 87%. They possess allergenic potential and are accepted as new isoallergens. One protein, designated as Dau c 1-like is >50% identical with the new isoallergens but exhibits no allergenicity. Sequence and structural comparisons of this protein with the known Dau c 1 isoallergens offer relevant clues about putative structural IgE epitopes.Identification of new isoallergens and the identification of IgE epitopes may contribute to a more refined component resolved diagnosis and may lay ground for further epitope mapping and personalized targeted treatment approaches of carrot allergy in preclinical and clinical studies.