In double-blind, placebo-controlled, oral food challenges with fish, a 12-fold higher false-negative rate was found compared with other food antigens. In an effort to elucidate this discrepancy, cooked lyophilized fish extracts (used in double-blind, placebo-controlled, oral food challenges) were compared with cooked, nonlyophilized fish extracts (used in open challenges) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoblot, and ELISA-inhibition assays. Altered fish allergenicity as a result of food processing was examined with canned tuna and salmon. Forty-five children and young adults with food allergies, including 18 patients with IgE-mediated hypersensitivity to fish, were challenged with canned tuna. All 45 challenges with canned tuna were negative. Two of these patients are allergic to salmon and also have negative reactions to challenges with canned salmon. In vitro investigation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of tuna and salmon extracts revealed a striking loss of definable protein fractions in the canned fish extract when compared with raw and cooked fish extracts, and immunoblot analyses demonstrated minimal IgE-specific binding to the canned fish extracts. In addition, decreased allergenicity of the canned tuna and salmon was demonstrated by ELISA-inhibition assay and by negative oral challenges with canned salmon in two patients allergic to salmon. Collectively, these findings suggest that some of the major allergens responsible for IgE-mediated food allergy to fish are more labile than previously recognized.