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Background: Delayed food anaphylaxis upon consumption of red meat is attributed to specific IgE-antibodies directed to galactose-α-1,3-galactose (α-Gal). Anaphylactic reactions may occur after ingestion of meat from different mammals, mainly beef and pork, but reactions to lamb, rabbit or horse have also been reported. In particular, pork kidney has been shown to trigger symptoms that were more severe and occurred within a shorter delay. The objective of the present study was the identification and characterization of pork kidney proteins carrying α-Gal carbohydrates and mediating delayed allergic reactions through specific IgE to α-Gal. Materials and methods: A cohort of 59 patients with specific IgE to α-Gal was screened by immunoblot for IgE-reactive proteins in pork kidney extract. Proteins were purified by affinity chromatography and identified by Edman sequencing and peptide mass fingerprinting. Isolated proteins were used in immunoassays using patient sera and α-Gal specific antibodies. Allergenicity was assayed in basophil activation and skin prick test. Results: Multiple IgE-binding proteins were detected in protein extracts of pork kidney by immunoblot using patient sera and an anti-α-Gal antibody. Reactive bands were located in the high molecular weight range of 100 to ≥200 kDa. Two major IgE-binding proteins were identified as porcine angiotensin I converting enzyme (ACE I) and aminopeptidase N (AP-N). IgE-binding to both proteins was lost by periodate treatment, resulting in oxidation of carbohydrates. Addition of α-Gal inhibited IgE-reactivity to both peptidases. Allergenicity was confirmed by activation of patient basophils and positive skin prick tests. Conclusions: Two IgE-reactive cell membrane peptidases carrying α-Gal epitopes were identified in pork kidney, a tissue which is known as potent inducer of red meat-induced anaphylaxis. Allergenicity and clinical relevance of these proteins were confirmed in patients with delayed anaphylaxis to red meat by skin prick test and basophil activation.

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