Allergy to fish is one of the most common food allergies. Gad c 1 is the only fish allergen which has been purified and characterized. Other allergens have been detected by Western blot in cod extracts. We have now improved the Western-blot procedure in order to characterize fish IgE-reactive proteins from extracts prepared under different conditions: pre-rigor mortis and post-rigor mortis, EDTA addition or not, and DEAE ion-exchange chromatography. Several IgE-reactive protein bands have been identified over a wide molecular-weight range. In particular, the 104- and 130-kDa IgE-reactive protein bands were detected. These new bands may correspond to aggregates, as EDTA increased the relative amount of the 60-, 67-, 104-, and 130-kDa IgE-reactive protein bands in Western blot. All these bands were also detected by antiparvalbumin monoclonal antibody, specific to the first calcium-binding site. The longer period of storage increased the relative amounts of the 41-, 80-, 104-, and 130-kDa IgE-reactive protein bands. The 18-kDa band was detected only in fish stored for several days. In conclusion, we have described IgE-reactive protein bands over a wide molecular-weight range (12-130 kDa) in Western blot of cod extract, and shown that EDTA and storage conditions may influence the relative distribution of IgE-reactive protein bands.