Person: Pastor Vargas, Carlos
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First Name
Carlos
Last Name
Pastor Vargas
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Ciencias Químicas
Department
Bioquímica y Biología Molecular
Area
Bioquímica y Biología Molecular
Identifiers
14 results
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Now showing 1 - 10 of 14
Item Novel liquid chromatography–mass spectrometry method for sensitive determination of the mustard allergen Sin a 1 in food(Food Chemistry, 2015) Maria Posada-Ayala; Álvarez Llamas, Gloria; Aroa S. Maroto; Xavier Maes; Esther Muñoz-Garcia; Villalba Díaz, María Teresa; Rodríguez García, Rosalía; Marina Perez-Gordo; Vivanco Martínez, Fernando; Pastor Vargas, Carlos; Javier Cuesta-HerranzMustard is a condiment added to a variety of foodstuffs and a frequent cause of food allergy. A new strategy for the detection of mustard allergen in food products is presented. The methodology is based on liquid chromatography analysis coupled to mass spectrometry. Mustard allergen Sin a 1 was purified from yellow mustard seeds. Sin a 1 was detected with a total of five peptides showing a linear response (lowest LOD was 5ng). Sin a 1 was detected in mustard sauces and salty biscuit (19±3mg/kg) where mustard content is not specified. Sin a 1, used as an internal standard, allowed quantification of this mustard allergen in foods. A novel LC/MS/MS SRM-based method has been developed to detect and quantify the presence of mustard. This method could help to detect mustard allergen Sin a 1 in processed foods and protect mustard-allergic consumers.Item Sensitive detection of major food allergens in breast milk: first gateway for allergenic contact during breastfeeding(Allergy, 2015) Pastor Vargas, Carlos; Maroto, Aroa; Díaz‐Perales, Araceli; Villalba Díaz, María Teresa; Casillas Diaz, Natalia; Vivanco Martínez, Fernando; Cuesta‐Herranz, JavierFood allergy is recognized as a major public health issue, especially in early childhood. It has been hypothesized that early sensitization to food allergens maybe due to their ingestion as components dissolved in the milk during the breastfeeding, explaining reaction to a food, which has never been taken before. Thus, the aim of this work has been to detect the presence of the food allergens in breast milk by microarray technology. We produced a homemade microarray with antibodies produced against major food allergens. The antibody microarray was incubated with breast milk from 14 women collected from Fundación Jiménez Díaz Hospital. In this way, we demonstrated the presence of major foods allergens in breast milk. The analysis of allergens presented in breast milk could be a useful tool in allergy prevention and could provide us a key data on the role of this feeding in tolerance induction or sensitization in children.Item Identification of vitellogenin as an allergen in Beluga caviar allergy(Allergy, 2008) Pérez‐Gordo, M.; Sánchez‐García, S.; Cases, B.; Cuesta‐Herranz, J.; Pastor Vargas, Carlos; Vivanco Martínez, FernandoItem Watermelon Profilin: Characterization of a Major Allergen as a Model for Plant-Derived Food Profilins(International Archives of Allergy Immunology, 2010) Cases, Bárbara ; Pastor Vargas, Carlos; Gil Dones, Félix; Perez-Gordo, Marina; Maroto, Aroa; las Heras, Manuel de las; Vivanco Martínez, Fernando; Cuesta-Herranz, JavierBackground: Plant profilins have been reported as minor allergens. They are a well-known pan-allergen family responsible for cross-reactivity between plant-derived foods and pollens. Watermelon profilin has been reported to be a major allergen in watermelon (Citrullus lanatus).The aim of this study was to characterize recombinant watermelon profilin, confirming its reactivity for diagnostic purposes and the development of immunotherapy. Methods: Native profilin was purified from watermelon extract by affinity chromatography using poly-L-proline. Recombinant His-tagged profilin was produced in Pichia pastoris yeast using pPICZαA vector and purified by metal chelate affinity chromatography. ELISA and immunoblot were carried out with sera from 17 watermelon-allergic patients. Biological activity was tested by the basophil activation test. Results: Native profilin and recombinant profilin were purified and identified by mass spectrometry. Both show similar IgE reactivity in vitro and are biologically active. Conclusions: Similarities were found in the IgE-binding patterns and biological activity of recombinant profilin and native profilin. Recombinant profilin may be a powerful tool for specific diagnosis.Item Targeting antigens to an invariant epitope of the MHC Class II DR molecule potentiates the immune response to subunit vaccines(Virus Research, 2011) Pérez-Filgueira, Mariano; Barderas, María G.; Alonso, Covadonga; José M, Escribano; Gil Dones, Félix; Pastor Vargas, Carlos; Vivanco Martínez, FernandoRecombinant subunit and peptidic vaccines in general present a reduced immunogenicity in vaccinated individuals with respect to the whole pathogen from which they derived. The generation of strong immune responses to these vaccines requires the use of potent adjuvants, high antigen doses and repetitive vaccinations. In this report, we document the enhanced antibody response obtained against two recombinant subunit vaccines by means of targeting to antigen-presenting cells by a recombinant single chain antibody. This antibody, named APCH1, recognizes an epitope of MHC Class II DR molecule preserved in different animal species, including humans. We showed that vaccinal antigens translationally fused to APCH1 antibody and produced by recombinant baculoviruses in insect larvae (Trichoplusia ni), elicited an increased antibody response in comparison with the same antigens alone or fused to a carrier molecule. These results suggest that targeting of antigens to this invariant MHC Class II epitope has immunopotentiating effects that could circumvent the reduced potency of peptidic or subunit vaccines, opening the possibility of widespread application of APCH1 as a new adjuvant antibody of general use.Item Allergy to kiwi: a double-blind, placebo-controlled food challenge study in patients from a birch-free area.(Journal of Allergy and Clinical Immunology, 2004) Alemán, Ana; Sastre, Joaquín; Quirce, Santiago; Heras González, Manuel De Las; Carnés, Jerónimo; Fernández-Caldas, Enrique; Pastor Vargas, Carlos; Blázquez, Ana Belén; Vivanco Martínez, Fernando; Cuesta-Herranz, Javierackground: Allergy to kiwi fruit is being increasingly reported, but it has never been evaluated by means of a double-blind, placebo-controlled food challenge (DBPCFC) study. Objective: We sought to assess kiwi allergy on the basis of a DBPCFC and identify the patterns of allergen recognition in sensitized patients from a birch-free area. Methods: Forty-three patients with allergy symptoms who were sensitized to kiwi were evaluated by means of clinical history, skin tests, IgE determinations, and DBPCFCs. The pattern of allergen recognition was assessed by means of IgE immunoblotting. Sequence analysis of IgE-binding bands was performed by using Edman degradation. Results: DBPCFCs were performed in 33 patients; 4 patients had experienced severe anaphylaxis, and 6 patients declined informed consent. DBPCFC results were positive in 23 patients and negative in 10 patients. The most frequent clinical manifestation was oral allergy syndrome. Twenty-one percent of the patients were not allergic to pollen. Forty-six percent of patients experienced systemic symptoms, and this happened with higher frequency in patients not allergic to pollen (100%). Twenty-eight percent of the patients were sensitized to latex. The IgE-binding bands in kiwi extract more frequently recognized by patient sera were those of 30, 24, 66, and 12 kd, and they could not be associated with any pattern of kiwi-induced allergic reactions. Conclusion: The results provide evidence that kiwi allergy is not a homogeneous disorder because several clinical subgroups can be established. No definite allergen-recognition pattern was associated with the type of allergic reactions to kiwi. One of 5 patients with kiwi allergy was not allergic to pollen, and these patients had the highest risk of systemic reactions to kiwi.Item Serine 132 Is the C3 Covalent Attachment Point on the CH1 Domain of Human IgG1(Journal of Biological Chemistry, 2001) Vidarte, Luis; Pastor Vargas, Carlos; Mas, Sebastian; Blázquez, Ana Belen; Rios, Vivian de los; Guerrero, Rosa; Vivanco Martínez, FernandoThe covalent binding of C3 (complement component C3) to antigen-antibody complexes (Ag.Ab; immune complexes (ICs)) is a key event in the uptake, transport, presentation, and elimination of Ag in the form of Ag.Ab.C3b (IC.C3b). Upon interaction of C3 with IgG.IC, C3b.C3b.IgG covalent complexes are formed that are detected on SDS-polyacrylamide gel electrophoresis by two bands corresponding to C3b.C3b (band A) and C3b.IgG (band B) covalent complexes. This allows one to evaluate the covalent binding of C3b to IgG antibodies. It has been described that C3b can attach to both the Fab (on the CH1 domain) and the Fc regions of IgG. Here the covalent interaction of C3b to the CH1 domain, a region previously described spanning residues 125-147, has been studied. This region of the CH1 domain is exposed to solvent and contains a cluster of six potential acceptor sites for ester bond formation with C3b (four Ser and two Thr). A set of 10 mutant Abs were generated with the putative acceptor residues substituted by Ala, and we studied their covalent interaction with C3b. Single (Ser-131, Ser-132, Ser-134, Thr-135, Ser-136, and Thr-139), double (positions 131-132), and multiple (positions 134-135-136, 131-132-134-135-136, and 131-132-134-135-136-139) mutants were produced. None of the mutants (single, double, or multiple) abolished completely the ability of IgG to bind C3b, indicating the presence of C3b binding regions other than in the CH1 domain. However, all mutant Abs, in which serine at position 132 was replaced by Ala, showed a significant decrease in the ability to form C3b.IgG covalent complexes, whereas the remaining mutants had normal activity. In addition we examined ICs using the F(ab')2 fragment of the mutant Abs, and only those containing Ala at position 132 (instead of Ser) failed to bind C3b. Thus Ser-132 is the binding site for C3b on the CH1 domain of the heavy chain, in the Fab region of human IgG.Item Identification of potential allergens involved in systemic reactions to melon and watermelon(Annals of allergy, asthma & immunology, 2010) González-Mancebo, Eloina; López-Torrejón, Gema; González de Olano, David; Cembellín Santos, Sara; Gandolfo-Cano, Mar; Meléndez, Amaya; Salcedo, Gabriel; Cuesta-Herranz, Javier; Vivanco Martínez, Fernando; Pastor Vargas, CarlosItem The covalent interaction of C3 with IgG immune complexes(Molecular Immunology, 1999) Vivanco Martínez, Fernando; Muñoz, Esther; Vidarte, Luis; Pastor Vargas, CarlosAntigens (Ags) are converted into immune complexes (antigen-antibody complexes, IC) as soon as they encounter their specific antibodies (Abs). In fluids containing complement, the process of IC formation and fixation of complement components occur simultaneously. Hence, the formation of Ag-Ab-complement complexes is the normal way of eliminating Ags from a host. C3b-C3b-IgG covalent complexes are immediately formed on interaction of serum C3 with IgG-IC. These C3b-C3b dimers constitute the core for the assembly of C3/C5-convertase on the IC, which are subsequently converted into iC3b-iC3b-IgG by the complement regulators. These complexes are detected on SDS-PAGE by two bands of molecular composition, C3alpha65-C3alpha43 (band A) and C3alpha65-heavy chain of the Ab (band B), which correspond to C3b-C3b and C3b-IgG covalent interaction respectively, and that identify opsonized IC (C3b-IC). C3b can attach to Fab and Fc regions of the Ab molecule with similar efficiency. The presence of multiple C3b binding regions on IgG is considered an advantageous characteristic that facilitates the elimination of Ags in the form of C3b(n)-IC. Ab molecules on the IC recognize the Ag, and also serve as a very good acceptor for C3b binding. In this way, Ags, even if they have no acceptor sites for C3b, can be efficiently processed and removed. When C3 is activated in serum by IC or other activators, secondary C3b-IgG covalent complexes are generated, with bystander monomeric circulating IgG, and thus constitute, physiological products of complement activation. These complexes gain importance when IgG concentration is extremely high as in cases of infusion of intravenous IgG (IVIG) in several pathologies. The covalent attachment of activated complement C3 (C3b, iC3b, C3 d,g) to Ags or IC links innate and adaptative immunity by targeting Ags to different cells of the immune system (follicular dendritic cells, phagocytes, B cells). Hence C3b marks Ags definitively, from the earliest contact with the innate immune system until their complete elimination from the host.Item Identification of Major Allergens in Watermelon(International Archives on Allergy & Immunology, 2009) Pastor Vargas, Carlos; Cuesta-Herranz, Javier; Cases, Barbara; Pérez-Gordo, Marina; Figueredo, Elena; Heras González, Manuel De Las; Vivanco Martínez, FernandoBackground: Watermelon is a worldwide consumed Cucurbitaceae fruit that can elicit allergic reactions. However, the major allergens of watermelon are not known. The aim of this study is to identify and characterize major allergens in watermelon. Methods: Twenty-three patients allergic to watermelon took part in the study. The diagnosis was based on a history of symptoms and positive skin prick-prick tests to watermelon, confirmed by positive open oral challenge testing to watermelon pulp. Allergenic components were detected by SDS-PAGE and immunoblotting. Molecular characterization of IgE-binding bands was performed by N-terminal amino acid sequencing and mass spectrometry. Allergens were purified combining several chromatographic steps. Results: Several IgE binding bands (8-120 kDa) were detected in watermelon extract. Three major allergens were identified as malate dehydrogenase (36 kDa), triose phosphate isomerase (28 kDa) and profilin (13 kDa). Purified allergens individually inhibited IgE binding to the whole watermelon extract. Conclusions: All in all these results indicate that malate dehydrogenase, triose phosphate isomerase and profilin are major allergens involved in watermelon allergy.