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Original Article


Expression of a Recombinant Protein Immunochemically Equivalent to the Major Anisakis simplex allergen Ani s 1


I Ibarrola,1* MC Arilla,1* MD Herrero,2 MI Esteban,2 A Martínez,1 JA Asturias1

1Bial-Arístegui, R&D Department, Bilbao, Spain
2 Allergy Service, General Hospital, Segovia, Spain

J Investig Allergol Clin Immunol 2008; Vol. 18(2): 78-83



Background: Anisakis simplex is a nematode which can parasitize humans, producing anisakiasis and can induce immunoglobulin-(Ig)- E–mediated allergic symptoms. Parasite recombinant proteins, such as the major allergen Ani s 1, may be useful tools to avoid misdiagnosis of A simplex allergy due to cross-reactivity when whole parasite extracts are used.

Objective: To obtain Ani s 1 allergen as a recombinant protein with IgE-binding properties similar to its natural counterpart.

Methods: Ani s 1-encoding cDNA was amplifi ed by polymerase chain reaction and cloned. The allergen was expressed in Escherichia coli as a nonfusion protein. Natural and recombinant Ani s 1 were investigated by means of Western blotting, enzyme allergosorbent test, enzyme-linked immunosorbent assay (ELISA), and ELISA inhibition using sera from 53 patients with A simplex allergy.

Results: Residues of the amino acid sequence of the encoded protein were 99.4% identical to the reported one. Purifi ed rAni s 1 was obtained with a yield of 2 mg/L of culture while the yield of the natural counterpart was only 50 μ/g of larvae. rAni s 1 reactivity was not significantly different from that of the natural allergen; the correlation was excellent (ρ = 0.92, P < .001). ELISA-inhibition experiments showed that the dose-response inhibition curve obtained with rAni s 1 overlapped with that of nAni s 1. In an enzyme allergosorbent analysis, 86.8% of the A simplex-allergic patient sera reacted to rAni s 1.

Conclusion: Recombinant Ani s 1 is immunochemically equivalent to its natural counterpart and therefore might be useful for the in vitro diagnosis of anisakiasis and A simplex-mediated allergy.

Key words: Nematode. Recombinant allergen. Biologically active allergen Diagnostic tool. Major allergen.