An antibody-based ELISA for quantification of Ani s 1, a major allergen from Anisakis simplex.
Anisakis simplex is a nematode parasite that can infect humans who have eaten raw or undercooked seafood. Larvae invading the gastrointestinal mucosa excrete/secrete proteins that are implicated in the pathogenesis of anisakiasis and can induce IgE-mediated symptoms. Since Ani s 1 is a potent secreted allergen with important clinical relevance, its measurement could assess the quality of allergenic products used in diagnosis/immunotherapy of Anisakis allergy and track the presence of A. simplex parasites in fish foodstuffs. An antibody-based ELISA for quantification of Ani s 1 has been developed based on monoclonal antibody 4F2 as capture antibody and biotin-labelled polyclonal antibodies against Ani s 1 as detection reagent. The dose-response standard curves, obtained with natural and recombinant antigens, ranged from 4 to 2000 ng/ml and were identical and parallel to that of the A. simplex extract. The linear portion of the dose-response curve with nAni s 1 was between 15 and 250 ng/ml with inter-assay and intra-assays coefficients of variation less than 20% and 10%, respectively. The assay was specific since there was no cross-reaction with other extracts (except Ascaris extracts) and was highly sensitive (detection limit of 1.8 ng/ml), being able to detect Ani s 1 in fish extracts from codfish and monkfish.
Arilla
M C
MC
Research & Development Department, Bial-Arístegui, Alameda Urquijo, 27, 48008 Bilbao, Spain.
Ibarrola
I
I
Martínez
A
A
Monteseirín
J
J
Conde
J
J
Asturias
J A
JA
D000485:Allergens; D000818:Animals; D017129:Anisakiasis / Q000175:diagnosis; D017163:Anisakis / Q000276:immunology*; D000909:Antibodies, Helminth / Q000276:immunology; D002135:Calcium-Binding Proteins; D045744:Cell Line, Tumor; D003445:Crustacea; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005393:Fish Diseases / Q000175:diagnosis; D005399:Fishes / Q000469:parasitology; D015801:Helminth Proteins; D006825:Hybridomas; D051379:Mice; D008807:Mice, Inbred BALB C; D008974:Mollusca; D012680:Sensitivity and Specificity; D013154:Spleen / Q000166:cytology
D000485:Allergens; D000909:Antibodies, Helminth; D002135:Calcium-Binding Proteins; D015801:Helminth Proteins; C412511:troponin-like protein, Anisakis simplex