RESUMEN
BACKGROUND: Characterized native and recombinant Hevea brasiliensis (rHev b) natural rubber latex (NRL) allergens are available to assess patient allergen sensitization profiles. OBJECTIVE: Quantification of individual IgE responses to the spectrum of documented NRL allergens and evaluation of cross-reactive carbohydrate determinants (CCDs) for more definitive diagnosis. METHODS: Sera of 104 healthcare workers (HCW; 51 German, 21 Portuguese, 32 American), 31 spina bifida patients (SB; 11 German, 20 Portuguese) and 10 Portuguese with multiple surgeries (MS) were analysed for allergen-specific IgE antibody (sIgE) to NRL, single Hev b allergens and CCDs with ImmunoCAP technology. RESULTS: In all patient groups rHev b 5-sIgE concentrations were the most pronounced. Hev b 2, 5, 6.01 and 13 were identified as the major allergens in HCW and combined with Hev b 1 and Hev b 3 in SB. In MS Hev b 1 displayed an intermediate relevance. Different sIgE antibody levels to native Hevea brasiliensis (nHev b) 2 and rHev b 6.01 allowed discrimination of SB with clinical relevant latex allergy vs. those with latex sensitization. Sensitization profiles of German, Portuguese and American patients were equivalent. rHev b 5, 6.01 and nHev b 13 combined detected 100% of the latex-allergic HCW and 80.1% of the SB. Only 8.3% of the sera showed sIgE response to CCDs. CONCLUSIONS: Hev b 1, 2, 5, 6.01 and 13 were identified as the major Hev b allergens and they should be present in standardized latex extracts and in vitro allergosorbents. CCDs are only of minor relevance in patients with clinical relevant latex allergy. Component-resolved diagnostic analyses for latex allergy set the stage for an allergen-directed immunotherapy strategy.
Asunto(s)
Reacciones Antígeno-Anticuerpo/inmunología , Inmunoglobulina E/inmunología , Hipersensibilidad al Látex/diagnóstico , Goma , Adolescente , Adulto , Antígenos de Plantas/biosíntesis , Antígenos de Plantas/genética , Antígenos de Plantas/inmunología , Carbohidratos/inmunología , Niño , Preescolar , Reacciones Cruzadas/inmunología , Epítopos/inmunología , Femenino , Alemania , Personal de Salud , Hevea/química , Hevea/genética , Humanos , Inmunoglobulina E/sangre , Hipersensibilidad al Látex/sangre , Hipersensibilidad al Látex/inmunología , Masculino , Persona de Mediana Edad , Portugal , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/inmunología , Disrafia Espinal/complicaciones , Estados UnidosRESUMEN
BACKGROUND: The latex of Hevea brasiliensis trees contains a complex proteome that includes a range of allergenic proteins. Current latex extracts that are used for the diagnosis of latex allergy still lack important allergens. We aimed to devise a production process for an improved reagent that would ideally contain the complete latex allergome. METHODS: Latex C-serum was fractionated by ammonium sulfate precipitation, and B- and C-serum proteins were then separated by anion exchange chromatography. Proteins eluting within defined salt concentration ranges were pooled into six final fractions. Fractions were evaluated by two-dimensional electrophoresis and subsequent IgE immunoblot for their spectrum of allergens. The presence of the most important latex allergens in the fractions was checked by Western blot analyses. Each fraction was further evaluated by skin prick test (SPT). RESULTS: Reproducibility of the preparation method was demonstrated with two batches of latex. Comparison of latex B- and C-serum to the six fractions showed a remarkable increase in the number of detectable allergens in the fractions. The presence of the latex allergens Hev b 1-8 and Hev b 13 in the fractions was demonstrated. In SPTs, the fractions produced wheal-and-flare reactions comparable to commercial latex extracts. CONCLUSIONS: This method provides reproducible latex protein fractions of high allergen content for the diagnosis of latex allergy.
Asunto(s)
Alérgenos/metabolismo , Antígenos de Plantas/metabolismo , Hevea/inmunología , Hipersensibilidad al Látex/diagnóstico , Látex/inmunología , Látex/metabolismo , Proteínas de Plantas/metabolismo , Alérgenos/inmunología , Antígenos de Plantas/inmunología , Antígenos de Plantas/aislamiento & purificación , Humanos , Látex/aislamiento & purificación , Hipersensibilidad al Látex/inmunología , Proteínas de Plantas/inmunología , Pruebas CutáneasRESUMEN
BACKGROUND: Hev b 7 is a Hevea brasiliensis latex allergen with sequence identities of 39% to 42% to patatins recently identified as potato allergens. The complementary DNAs encoding 2 different Hev b 7 isoforms were previously reported. OBJECTIVE: The aim of this study was to determine the sequence variation of Hev b 7 and to compare the IgE reactivity of individual isoforms in vitro and in vivo. A further objective was to evaluate possible cross-reactivities between Hev b 7 and patatins and proteins from banana and avocado. METHODS: An H brasiliensis lambda ZAP complementary DNA (cDNA) library was screened with use of a Hev b 7 cDNA probe. Four Hev b 7 isoforms were produced in recombinant form and their IgE-binding capacities were compared. IgE immunoblot inhibitions and ELISA inhibition assays were used to investigate the possible cross-reactivity between Hev b 7 and recombinant potato patatin and proteins from avocado and banana. RESULTS: Two new isoforms, S2 and D2, were identified by sequencing 32 cDNA clones with full-length coding regions. All 4 recombinant isoforms displayed esterase activity and identical IgE-binding capacities. The new isoforms S2 and D2 were evaluated in skin prick tests and provoked responses equivalent to natural Hev b 7. No cross-reactivity was observed between Hev b 7 isoforms and potato patatin and proteins from avocado and banana. CONCLUSIONS: All 4 recombinant Hev b 7 isoforms have equivalent IgE-binding capacity and therefore represent suitable reagents for the development of in vitro and in vivo diagnostic tests. Hev b 7, patatins, and their homologs appear not to contribute to cross-reactivity in the latex-fruit syndrome.