title: Expression of a recombinant protein immunochemically equivalent to the major Anisakis simplex allergen Ani s 1.
abstract: 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 amplified 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. Purified rAni s 1 was obtained with a yield of 2 mg/L of culture while the yield of the natural counterpart was only 50 micro/g of larvae. rAni s 1 reactivity was not significantly different from that of the natural allergen; the correlation was excellent (p = 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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Bial-Arístegui, R&D Department, Bilbao, Spain
authors.forename: I
authors.initials: I
authors.lastname: Ibarrola
authors.forename: M C
authors.initials: MC
authors.lastname: Arilla
authors.forename: M D
authors.initials: MD
authors.lastname: Herrero
authors.forename: M I
authors.initials: MI
authors.lastname: Esteban
authors.forename: A
authors.initials: A
authors.lastname: Martínez
authors.forename: J A
authors.initials: JA
authors.lastname: Asturias
mesh_terms: D000485:Allergens / Q000096:biosynthesis* / Q000235:genetics* / Q000276:immunology; D000818:Animals; D017129:Anisakiasis / Q000097:blood / Q000175:diagnosis* / Q000276:immunology; D017163:Anisakis; D000909:Antibodies, Helminth / Q000097:blood / Q000235:genetics / Q000276:immunology*; D000947:Antigens, Helminth / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology*; D002135:Calcium-Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D018509:DNA, Helminth / Q000235:genetics / Q000276:immunology; D004926:Escherichia coli; D015801:Helminth Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D006801:Humans; D006969:Hypersensitivity, Immediate / Q000175:diagnosis / Q000469:parasitology; D007120:Immunochemistry; D007073:Immunoglobulin E / Q000097:blood / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology*
chemical_list: D000485:Allergens; D000909:Antibodies, Helminth; D000947:Antigens, Helminth; D002135:Calcium-Binding Proteins; D018509:DNA, Helminth; D015801:Helminth Proteins; D011994:Recombinant Proteins; C412511:troponin-like protein, Anisakis simplex; D007073:Immunoglobulin E