Immunoblot Inhibition Studies Research Articles

Identification of a vicilin-like major allergen from Prosopis juliflora exhibiting cross- reactivity with legume food allergens

BackgroundProsopis juliflora is a clinically relevant allergic sensitizer worldwide and shares cross-reactivity with allergens from several tree pollen and food. The present study aims to purify and immunobiochemically characterize a major allergen from Prosopis pollen. The allergen was further investigated for its cross-reactivity with legume allergens. MethodsProsopis extract was fractionated by Q Sepharose and Superdex 75 gel filtration column to purify the allergen. Specific IgE against purified protein was estimated via ELISA and immunoblot. The protein was subjected to mass spectrometric analysis. Glycan characterization was performed by Schiff staining and lectin binding assay followed by deglycosylation studies. The functional activity of the purified protein was evaluated by the basophil activation test. Cross-reactivity was assessed by inhibition studies with legume extracts. ResultsA 35 kDa protein was purified and showed 75% IgE reactivity with the patients’ sera by ELISA and immunoblot. Glycan characterization of protein demonstrated the presence of terminal glucose and mannose residues. A reduction of 40% and 27% in IgE binding was observed upon chemical and enzymatic deglycosylation of the protein, respectively. The glycoprotein allergen upregulates the expression of CD203c on basophils which was significantly reduced upon deglycosylation, signifying its biological ability to activate the effector cells. The identified protein shared significant homology with Lup an 1 from the lupine bean. Immunoblot inhibition studies of the purified allergen with legume extracts underlined high cross-reactive potential. Complete inhibition was observed with peanut and common bean, while up to 70% inhibition was demonstrated with soy, black gram, chickpea, and lima bean. ConclusionA 35 kDa vicilin-like major allergen was isolated from P. juliflora. The protein possesses glycan moieties crucial for IgE binding and basophil activation. Furthermore, the purified protein shows homology with Lup an 1 and exhibits cross-reactivity with common edible legume proteins.

Mite allergoids coupled to nonoxidized mannan from Saccharomyces cerevisae efficiently target canine dendritic cells for novel allergy immunotherapy in veterinary medicine

We have recently reported that grass pollen allergoids conjugated with nonoxidized mannan of Saccharomyces cerevisae using glutaraldehyde results in a novel hypoallergenic mannan-allergen complex with improved properties for allergen vaccination. Using this approach, human dendritic cells show a better allergen uptake and cytokine profile production (higher IL-10/IL-4 ratio) for therapeutic purposes. Here we aim to address whether a similar approach can be extended to dogs using canine dendritic cells. Six healthy Spanish Greyhound dogs were used as blood donors to obtain canine dendritic cells (DC) derived from peripheral blood monocytes. Allergens from Dermatophagoides farinae mite were polymerized and conjugated with nonoxidized mannan. Nuclear magnetic resonance (NMR), gel electrophoresis (SDS-PAGE), immunoblotting and IgE-ELISA inhibition studies were conducted to evaluate the main characteristics of the allergoid obtained. Mannan-allergen conjugate and controls were assayed in vitro for canine DC uptake and production of IL-4 and IL-10. The results indicate that the conjugation of D. farinae allergens with nonoxidized mannan was feasible using glutaraldehyde. The resulting product was a polymerized structure showing a high molecular weight as detected by NMR and SDS-PAGE analysis. The mannan-allergen conjugate was hypoallergenic with a reduced reactivity with specific dog IgE. An increase in both allergen uptake and IL-10/IL-4 ratio was obtained when canine DCs were incubated with the mannan-allergen conjugate, as compared with the control allergen preparations (unmodified D. farinae allergens and oxidized mannan-allergen conjugate). We conclude that hypoallergenic D. farinae allergens coupled to nonoxidized mannan is a novel allergen preparation suitable for canine allergy immunotherapy targeting dendritic cells.

La normalización en ciencias de laboratorio clínico

Chickpea has become one of the most abundant crops consumed in the Mediterranean and also in western world. Chickpea allergy is reported in specific geographic areas and is associated with lentil and/or pea allergy. We investigated cross-reactivity between chickpea and pea/lentil/soybean/hazelnut. The IgE-binding profiles of chickpea globulin and pea/lentil/soybean/hazelnut extracts were analyzed by immunoblotting and immunoblot-inhibition studies. Inhibition-assay with pea/lentil completely suppressed IgE-binding to chickpea globulin allergens, while not so in the reciprocal inhibition. Pre-absorption of sera with chickpea globulin caused the disappearance of IgE-binding to protein on an immunoblot of soybean/hazelnut protein extract. These results suggest that cross-reactivity exists between chickpea and pea/lentil/soybean/hazelnut. Chickpea allergy is associated with lentil and/or pea allergy, but evidently may not present independently. This, together with the described asymmetric cross-reactivity and phylogenetic aspects, suggest that chickpea allergy is merely an expression of cross-reactivity, caused by pea and/or lentil as the “primary” allergen.

Cross-Reactivity Among Pho d 2 and Profilins from Different Pollen Allergenic Sources

RationaleProfilins are considered as an important panallergen being the IgE cross-reactivity among these proteins associated with multiple pollen sensitization. Sequence homology among pollen profilins is very high, ranging 67.4-89.5%. The objective was to evaluate the cross-reactivity among profilin from palm tree pollen (Phoenix dactylifera) and other pollen allergenic sources where profilin has been described.MethodsPalm tree profilin (Pho d 2) was purified by affinity chromatography in a poly-L-proline column, sequenced by LC/MS-MS and used to produce rabbit polyclonal antibodies. Identification of profilin in eleven pollen extracts (Ambrosia artemisiifolia, Artemisia vulgaris, Betula alba, Chenopodium album, Cynodon dactylon, Helianthus annus, Mercurialis perennis, Olea europaea, Parietaria judaica, Phleum pratense and Plantago lanceolata) was carried out by IgG ELISA and immunoblot using the polyclonal antibodies. Cross-reactivity studies were performed by IgG immunoblot inhibition assays, using the purified profilin for inhibition.ResultsHighly purified profilin (>95%) from palm tree pollen (Pho d 2) with a molecular weight of 14 kDa was obtained. Identity and purity of profilin was confirmed by LC/MS-MS. Presence of profilin was confirmed in all the extracts, except Chenopodium, by direct ELISA (O.D.>0.1). Identification of profilin by IgG immunoblot, and cross-reactivity with Pho d 2 by immunoblot-inhibition, was observed in Ambrosia, Betula, Cynodon, Helianthus, Mercurialis, Plantago and Phleum extracts.ConclusionsCross-reactivity among Pho d 2 and other profilins was demonstrated in Ambrosia, Betula, Cynodon, Helianthus, Mercurialis, Plantago and Phleum extracts by immunoblot inhibition studies. Purified Pho d 2 could be used for diagnosis and treatment of pollen profilin sensitization. RationaleProfilins are considered as an important panallergen being the IgE cross-reactivity among these proteins associated with multiple pollen sensitization. Sequence homology among pollen profilins is very high, ranging 67.4-89.5%. The objective was to evaluate the cross-reactivity among profilin from palm tree pollen (Phoenix dactylifera) and other pollen allergenic sources where profilin has been described. Profilins are considered as an important panallergen being the IgE cross-reactivity among these proteins associated with multiple pollen sensitization. Sequence homology among pollen profilins is very high, ranging 67.4-89.5%. The objective was to evaluate the cross-reactivity among profilin from palm tree pollen (Phoenix dactylifera) and other pollen allergenic sources where profilin has been described. MethodsPalm tree profilin (Pho d 2) was purified by affinity chromatography in a poly-L-proline column, sequenced by LC/MS-MS and used to produce rabbit polyclonal antibodies. Identification of profilin in eleven pollen extracts (Ambrosia artemisiifolia, Artemisia vulgaris, Betula alba, Chenopodium album, Cynodon dactylon, Helianthus annus, Mercurialis perennis, Olea europaea, Parietaria judaica, Phleum pratense and Plantago lanceolata) was carried out by IgG ELISA and immunoblot using the polyclonal antibodies. Cross-reactivity studies were performed by IgG immunoblot inhibition assays, using the purified profilin for inhibition. Palm tree profilin (Pho d 2) was purified by affinity chromatography in a poly-L-proline column, sequenced by LC/MS-MS and used to produce rabbit polyclonal antibodies. Identification of profilin in eleven pollen extracts (Ambrosia artemisiifolia, Artemisia vulgaris, Betula alba, Chenopodium album, Cynodon dactylon, Helianthus annus, Mercurialis perennis, Olea europaea, Parietaria judaica, Phleum pratense and Plantago lanceolata) was carried out by IgG ELISA and immunoblot using the polyclonal antibodies. Cross-reactivity studies were performed by IgG immunoblot inhibition assays, using the purified profilin for inhibition. ResultsHighly purified profilin (>95%) from palm tree pollen (Pho d 2) with a molecular weight of 14 kDa was obtained. Identity and purity of profilin was confirmed by LC/MS-MS. Presence of profilin was confirmed in all the extracts, except Chenopodium, by direct ELISA (O.D.>0.1). Identification of profilin by IgG immunoblot, and cross-reactivity with Pho d 2 by immunoblot-inhibition, was observed in Ambrosia, Betula, Cynodon, Helianthus, Mercurialis, Plantago and Phleum extracts. Highly purified profilin (>95%) from palm tree pollen (Pho d 2) with a molecular weight of 14 kDa was obtained. Identity and purity of profilin was confirmed by LC/MS-MS. Presence of profilin was confirmed in all the extracts, except Chenopodium, by direct ELISA (O.D.>0.1). Identification of profilin by IgG immunoblot, and cross-reactivity with Pho d 2 by immunoblot-inhibition, was observed in Ambrosia, Betula, Cynodon, Helianthus, Mercurialis, Plantago and Phleum extracts. ConclusionsCross-reactivity among Pho d 2 and other profilins was demonstrated in Ambrosia, Betula, Cynodon, Helianthus, Mercurialis, Plantago and Phleum extracts by immunoblot inhibition studies. Purified Pho d 2 could be used for diagnosis and treatment of pollen profilin sensitization. Cross-reactivity among Pho d 2 and other profilins was demonstrated in Ambrosia, Betula, Cynodon, Helianthus, Mercurialis, Plantago and Phleum extracts by immunoblot inhibition studies. Purified Pho d 2 could be used for diagnosis and treatment of pollen profilin sensitization.

Allergological characterisation of the storage mite Acarus gracilis (Acari: Acaridae)

BackgroundStorage mites of the genus Acarus can be responsible for allergic sensitisation in domestic environments. Acarus gracilis is a frequent species in some geographical regions of the Iberian Peninsula. Since the allergenicity of this mite has not been described before, the objectives of this study were to characterise it immunologically, and to compare it with the closely related and more extensively studied species Acarus siro. MethodsExtracts from A. gracilis and A. siro cultures were characterised by Lowry, 1D and 2D-SDS and IEF. Zymogram, and determination of different enzymatic activities were performed. Skin prick solution of A. gracilis was tested in consecutive patients attending the Hospital of Mérida (Extremadura, Spain). Serum samples from eight individuals with positive skin prick test were collected. IgE determination, immunoblot and immunoblot-inhibition studies were performed. ResultsExtracts of both species showed a very similar protein and allergenic profile. Allergens at 14 and 17kDa were clearly recognised in both extracts by serum samples. Immunoblot-inhibition studies demonstrated that both extracts were totally inhibited by the opposite one. Enzymatic activity was similar in both cases with the most important differences being in kallikrein, serine protease and collagenase activities. ConclusionThe storage mite A. gracilis has a similar protein and allergen profile to A. siro and can induce allergic sensitisation. Due to the higher prevalence of this species respect to A. siro in some regions, more studies are needed to determine the clinical significance of sensitisation to this storage mite species.

The transaldolase, a novel allergen of Fusarium proliferatum, demonstrates IgE cross-reactivity with its human analogue.

Fusarium species are among airborne fungi and recognized as causative agents of human atopic disorders. However, Fusarium allergens have not been well characterized and the lack of information limits clinical diagnosis and treatment of fungal allergy. The purpose of this study is to identify and characterize important allergens of F. proliferatum. IgE-reacting F. proliferatum components were identified by immunoblot using serum samples from patients of respiratory atopic diseases. Characterization of allergens and determination of IgE cross-reactivity were performed by cDNA cloning, then homologous expression and immunoblot inhibition studies. We identified nine different F. proliferatum components that can be recognized by IgE antibodies in 17 (28%) of the 60 atopic sera tested. Components with molecular masses of about 43, 37.5 and 36.5 kDa with IgE-binding frequencies of about 88, 47 and 53%, respectively, were considered as important allergens of F. proliferatum. The 37.5 kDa IgE-binding component was putatively considered as a transaldolase protein of F. proliferatum. The full-length cDNA of F. proliferatum transaldolase was subsequently cloned. It encodes an open reading frame of 312 amino acids and has sequence identifies of 73 and 61%, respectively, with Cladosporium and human transaldolases. The purified recombinant F. proliferatum transaldolase can inhibit the IgE-binding against the 37.5 kDa component of F. proliferatum and the transaldolase allergen from Cladosporium cladosporioides. More importantly, the recombinant F. proliferatum transaldolase can inhibit IgE-binding against human transaldolase in a concentration-dependent manner. Thus, a novel and important F. proliferatum transaldolase allergen was identified. In addition to IgE cross-reactivity between the Fusarium and the Cladosporium transaldolase allergens, IgE cross-reactivity between the Fusarium and the human transaldolases also exists and might contribute to atopic manifestations in the absence of exogenous allergen exposure.

Food allergy and cross-reactivity-chickpea as a test case

Chickpea has become one of the most abundant crops consumed in the Mediterranean and also in western world. Chickpea allergy is reported in specific geographic areas and is associated with lentil and/or pea allergy. We investigated cross-reactivity between chickpea and pea/lentil/soybean/hazelnut. The IgE-binding profiles of chickpea globulin and pea/lentil/soybean/hazelnut extracts were analyzed by immunoblotting and immunoblot-inhibition studies. Inhibition-assay with pea/lentil completely suppressed IgE-binding to chickpea globulin allergens, while not so in the reciprocal inhibition. Pre-absorption of sera with chickpea globulin caused the disappearance of IgE-binding to protein on an immunoblot of soybean/hazelnut protein extract. These results suggest that cross-reactivity exists between chickpea and pea/lentil/soybean/hazelnut. Chickpea allergy is associated with lentil and/or pea allergy, but evidently may not present independently. This, together with the described asymmetric cross-reactivity and phylogenetic aspects, suggest that chickpea allergy is merely an expression of cross-reactivity, caused by pea and/or lentil as the “primary” allergen.

Seafood hypersensitivity in mite sensitized individuals: is tropomyosin the only responsible allergen?

Seafood allergy has been related to mite sensitization, mainly mediated by the muscle protein tropomyosin. To determine the correlation between seafood hypersensitivity and mite sensitization (Dermatophagoides pteronyssinus and Chortoglyphus arcuatus, a highly prevalent storage mite in Spain) and to investigate the implication of tropomyosin in cross-reactivity. Patients from Northwest Spain were divided into 2 groups. The mite-seafood group contained 30 allergic mite individuals with a clinical history of food hypersensitivity. The mite group contained 40 individuals with positive skin prick test results to D pteronyssinus and C arcuatus but negative seafood test results. Specific IgE (sIgE) to whole mite and shrimp extracts, mite tropomyosin (rDer p 10), and shrimp tropomyosin (rPen a 1) were determined in each serum sample. Allergenic profiles were analyzed by immunoblot. Cross-reactivity studies were investigated by enzyme-linked immunosorbent assay and immunoblot inhibition studies. In the mite-seafood group, 71% of patients had positive sIgE results to shrimp and 55% of them to shrimp tropomyosin. A strong correlation was found between sIgE to shrimp tropomyosin and mite tropomyosin. Positive correlation was observed between sIgE to shrimp tropomyosin and severity of symptoms. In the mite group, none of the 20% of patients with sIgE to shrimp tested positive to shrimp tropomyosin. In the immunoblot inhibition experiment, the shrimp extract was totally inhibited by mite extract. These data suggest that primary sensitization is related to mite sensitization. Tropomyosin does not seem to be the main allergen involved in mite-seafood sensitization in mite sensitized individuals. High levels of sIgE to tropomyosin seem to be related to severity of symptoms.

Allergenicity and cross‐reactivity of pine pollen

Pine pollen has long been considered a non-allergenic pollen. The large size of the grain and its low levels of proteins are the main reasons invoked to explain this low allergenicity. The aim of this study was to describe the main allergenic bands of Pinus radiata (PR) and its cross-reactivity with other pine species, other conifers and grass pollen. Sixty-five pine-pollen-allergic patients (51% also sensitized to grass pollen) were studied. Skin prick tests (SPT) to a battery of allergens including PR, Pinus pinea, Pinus sylvestris, Pinus nigra and Cupressus sempervirens pollens and specific IgE determination to PR and Pinus strobus were performed. IgE-immunoblotting to a PR extract and other pine pollens was also carried out. UniCAP inhibition and immunoblotting inhibition studies were performed to assess the cross-reactivity between different pollens. The SPTs were positive with all the pine pollen extracts tested in 69% of the patients. Specific IgE was positive to PR or P. strobus in 77% of the patients, and to Lolium perenne in 51%. Nine different allergenic bands were detected. The two main allergens were a 42 kDa band recognized by 85% of the patients and a band of approximately 6-8 kDa recognized by 40%. A high degree of cross-reactivity was observed between different pine pollen species, but not between pines and C. sempervirens pollen. A partial cross-reactivity could be seen between pine and grass pollens only in patients also sensitized to L. perenne. Pine pollen should be considered as a potential allergenic pollen especially where this pollen is abundant. The detection of a high number of patients that were monosensitized to pine pollen suggests the possibility of treating these patients with specific immunotherapy.

Evaluation of Molecular Basis of Cross Reactivity between Rye and Bermuda Grass Pollen Allergens

Allergenic cross reactivity between the members of the Pooids (Lolium perenne, Phleum pratense, and Poa pratensis) and Chloridoids (Cynodon dactylon and Paspalum notatum) is well established. Studies using crude extracts in the past have demonstrated limited cross reactivity between the Pooids and the Chloridoids suggesting separate diagnosis and therapy. However, little is known regarding the molecular basis for the limited cross reactivity observed between the 2 groups of grasses. The present study was undertaken to gain insights into the molecular basis of cross allergenicity between the major allergens from rye and Bermuda grass pollens. Immunoblot inhibition tests were carried out to determine the specificity of the proteins involved in cross reactivity. Crude pollen extract and bacterially expressed and purified recombinant Lol p 1and Lol p 5 from rye grass were subjected to cross inhibition experiments with crude and purified recombinant Cyn d 1 from Bermuda grass using sera from patients allergic to rye grass pollen. The immunoblot inhibition studies revealed a high degree of cross inhibition between the group 1 allergens. In contrast, a complete lack of inhibition was observed between Bermuda grass group 1 allergen rCyn d 1, and rye grass group 5 allergen rLol p 5. Crude rye grass extract strongly inhibited IgE reactivity to Bermuda grass, whereas crude Bermuda grass pollen extract showed a weaker inhibition. Our data suggests that a possible explanation for the limited cross reactivity between the Pooids and Chloridoids may, in part, be due to the absence of group 5 allergen from Chloridoid grasses. This approach of using purified proteins may be applied to better characterize the cross allergenicity patterns between different grass pollen allergens.

Allergenicity and cross‐reactivity of Senecio pollen: identification of novel allergens using the immunoproteomics approach

The genus Senecio is the largest genus of the family Asteraceae (Compositae). The allergenicity of Senecio has not been assessed previously. The aim of this study was to investigate the allergens of Senecio jacobea pollen and to determine their immunological characteristics and clinical relevance. Fifty patients with rhinoconjunctivitis and a positive skin prick test (SPT) to Senecio were recruited. The clinical relevance of this pollen was assessed by means of a nasal provocation test (NPT). Allergens were characterized by one-dimensional electrophoresis (SDS-PAGE) and two-dimensional gel electrophoresis and immunoblotting. Furthermore, characterization and identification of the allergens were performed by mass spectrometry (MS). In vitro inhibition tests were performed to evaluate cross-reactivity with other pollen. Three predominant allergens, both in the intensity of reaction and the frequency of recognition by human-allergic sera, were 59 (60%), 42 (50%) and 31 kDa (50%). The two-dimensional analysis allowed the identification of several allergens. One spot around 42 kDa was identified as a protein homologous to pectate lyase and three other spots were homologous to malate dehydrogenase by MS. S. jacobea proteins showed cross-reactivity with other proteins of the Asteraceae family and also with Parietaria judaica. This was demonstrated by immunoblotting and ELISA inhibition studies. S. jacobea constitute a newly discovered allergenic source. It shows cross-reactivity with other members of the Asteraceae plant family as well as with P. judaica.

Cross‐reactivity between pollen allergens from common Pooideae grasses and rye

SummaryIn Germany, specific pollen extracts are currently used for immunotherapy of rye (Secale cereale) allergy. Like most common grasses, S. cereale belongs to the Pooideae subfamily. Using sera from grass pollen‐allergic patients, immunoblotting and ELISA inhibition studies demonstrate that a five‐grass allergenic extract from Anthoxanthum odoratum, Dactylis glomerata, Lolium perenne, Poa pratensis and Phleum pratense inhibits over 90% of IgE binding to rye allergens. This result confirms the high degree of homology between allergens from S. cereale and common grass species. We conclude that a five‐grass pollen mixture is appropriate for desensitization of patients who are allergic to rye pollen without the need for additional rye pollen extracts.

Gräserpollenextrakte eignen sich auch zur Hyposensibilisierung einer Roggenpollenallergie

In Germany, specific immunotherapy of patients with allergy against rye pollens (Secale cereale) is frequently done with different grass pollen extracts. We reassess the practice of applying rye allergens as single extract or in an individual allergen mixture in the context of the availability of a 5-grass tablet vaccine as speciality based on Anthoxanthum odoratum, Dactylis glomerata, Lolium perenne, Poa pratensis, Phleum pratense. Phylogenetically, Secale cereale shows a strong similarity to the most frequent grasses in Northern and Middle Europe. This similarity applies also to the allergen profile. In 3 studies with 256 grass pollen-allergic patients, the sensitization against rye as well as against other in Northern or Middle Europe frequently found grasses could be shown in all patients. Using sera from 15 grass pollen-allergic patients, it could be shown by immunoblot and ELISA inhibition studies that a 5-grass extract inhibits > 90% of IgE binding to rye allergens. The high cross-immunogenicity between Secale cereale and common grasses is reflected in medical practice in many European countries. Patients with rye allergy are treated with grass pollen extracts. According to the high cross-reactivity between grasses and rye-specific immunotherapy of rye allergy seems to be feasible also with a grass pollen extract. Related to existing data this is independent whether the therapeutic extract includes a single grass, a mixture of 5 grasses or other mixtures.

Anaphylaxis to Hippobosca equina (louse fly)

IgE-mediated allergy to insects different from Hymenoptera species is seldom reported. To describe and study the case of a previously nonatopic man with an anaphylactic reaction (grade III, Müeller) caused by a bite from a louse fly (Hippobosca equina). In vivo (skin prick tests) and in vitro (enzyme-linked immunosorbent assay and sodium dodecyl sulfatepolyacrylamide gel electrophoresis immunoblotting) tests were used for diagnosis. Cross-reactivity between H. equina and different insects was investigated. Results of skin prick tests and serum specific IgE were positive to H. equina. Immunoblot inhibition studies identified common bands in H. equina, Apis mellífera, and Musca domestica, but this cross-reactivity did not affect a band of 16 or 15 kDa. This molecular weight is similar to that of phospholipase A2 in A mellifera venom. We describe a case of IgE-mediated allergy to H. equina in which specific molecular band proteins seemed to be responsible for the reaction.

Currant allergy and the Rosaceae-grass pollen allergy syndrome: a case report

Despite the increasing use of currants in culinary recipes, currant allergy has rarely been reported. To study a case of currant allergy and to explore cross-reactivity between grass pollen and Rosaceae family fruit allergens. Skin prick tests to pollen and skin prick-to-prick tests with currants and peach were performed. Specific IgE levels were determined using the CAP method. We prepared a protein extract of 0.1 mg/mL in phosphate-buffered saline using red currant in the presence of protease inhibitors. Immunoblot inhibition studies were performed to explore cross-reactivity between grass pollen and currant allergens. Skin prick test results were positive to Dactylis, arizonic, and olive pollens. Results of skin prick-to-prick tests with fresh red and black currants were negative and positive, respectively, to peach. The specific IgE level was 5.7 KU/L to red currant and 2.92 KU/L to peach (CAP). Western blot analysis with red currant extract revealed specific IgE protein bands of 37 and 26 kDa. Preincubation of sera with extracts from red currant and peach inhibited both IgE bands, and preincubation with Dactylis pollen inhibited the 37-kDa band only. We report a case of allergy to grass pollen with an oral allergy syndrome involving several fruits from 2 different families of the Rosidae subclass confirmed by in vitro tests. Inhibition studies demonstrated cross-reactivity between different fruits (currant and raspberry) from the Rosidae subclass and were incomplete with grass pollen allergens.

Occupational respiratory allergy to roe deer

Although scattered reports have been published on roe deer allergenicity, no systematic studies of allergenicity or possible cross-reactions have appeared. To describe 2 patients with occupational roe deer allergy, demonstrated by positive skin and conjunctival provocation test results, and to note cross-reactions to other animal (mainly cow) allergens. Two workers at animal rehabilitation centers were sensitized to roe deer. One patient had a history of rhinoconjunctivitis and the other a history of rhinoconjunctivitis and probable asthma. Both patients underwent skin testing with a standard battery of inhaled and epithelial allergens and with roe deer hair and dander extract and conjunctival provocation tests with roe deer hair extract. Immunodetection for IgE (both patients) and IgE immunoblot inhibition tests to determine inhibitory effect (1 patient) were also performed. The results of skin tests and conjunctival provocation tests showed that both patients were sensitized to roe deer allergens. In one patient specific IgE to roe deer extract was detected, and this extract completely inhibited IgE binding to cow hair and dander extract in immunoblotting tests. Specific IgE to roe deer proteins could not be demonstrated in the other patient. Our results suggest that roe deer epidermal derivatives can cause occupational respiratory disease in exposed workers and that allergy to this species should be considered in individuals who present with similar symptoms and exposure histories. Immunoblot inhibition studies suggested the possibility of cross-reaction between roe deer proteins and cow proteins.

Evaluation of Cross-Reactivity between Holoptelea integrifolia and Parietaria judaica

Background:Holoptelea integrifolia and Parietaria judaica belong to the family Urticaceae, but are geographically distantly located. H. integrifolia is an important pollen allergen of India and sensitizes almost 10% of the atopic population in Delhi. P. judaica, on the other hand, is a very dominant pollen allergen of the Mediterranean region, sensitizing almost 80% of the allergic population. Since both these important pollen allergens belong to the family Urticaceae, the objective of the present study was to assess cross-reactivity between these two pollen allergens from different geographical regions. Methods: Cross-reactivity between these two pollen allergens was assessed on the basis of skin prick tests and ELISA, ELISA inhibition and immunoblot inhibition studies. Results: Out of 44 atopic Indian patients skin prick tested with H. integrifolia extract, 34% were found to be sensitized. All the patients sensitized to H. integrifolia also showed varying degrees of skin positivity to P. judaica pollen extract. ELISA and ELISA inhibition studies suggested strong cross-reactivity between H. integrifolia and P. judaica pollen. Immunoblot inhibition studies revealed that 14-, 16-, 28-, 38-, 42- and 46-kDa proteins are the cross-reactive proteins in H. integrifolia and P. judaica. However, Par j 1, the major allergen of P. judaica, is absent in H. integrifolia pollen. Conclusion:H. integrifolia and P. judaica pollens share cross-reactive as well as unique epitopes. The major allergen of P. judaica, Par j 1, seems to be absent in H. integrifolia pollen allergen.

Allergenic Components of Indian Jujube (Zizyphus mauritiana) Show IgE Cross-Reactivity with Latex Allergen

Background: ‘Latex-fruit syndrome’ has been well documented. A prevalence of latex allergy among medical workers of 6.8–8.6% had been reported in Taiwan. However, there has been no study to determine the importance and type of fruit hypersensitivity in latex-allergic patients in Taiwan. This study aimed to identify the allergenic components of Indian jujube (Zizyphus mauritiana) and characterize the cross-reactivity of specific IgE antibodies to latex allergen. Methods: Crude extracts were prepared from Indian jujube and from ammoniated natural rubber latex, and six medical workers and one patient with a history of fruit allergy underwent skin testing with routine allergens, latex, Indian jujube and other fruits. Sera from two Indian jujube skin test-positive latex-allergic subjects were used for allergen-specific IgE, immunoblotting, immunoblot inhibition and enzyme-linked immunosorbent assay (ELISA) inhibition studies. Results: Both patients had positive skin test responses and specific IgE assays to Indian jujube and latex extracts. Immunoblotting revealed that IgE from both subjects bound to a 42-kD latex protein and a 42-kD Indian jujube protein. In addition, IgE from one subject bound to a 30-kD Indian jujube protein. Preincubation of atopic sera with Indian jujube or latex extract demonstrated absent and/or marked inhibition of IgE binding. Moreover, anti-Indian jujube protein antibody-based ELISA was able to detect latex extracts. Conclusions: Our results add to findings regarding the ‘latex-fruit syndrome’ described in the literature, and further study of the cross-reacting allergens identified in Indian jujube may help to elucidate the mechanisms underlying this syndrome.

Characterization of allergens secreted by Anisakis simplex parasite: clinical relevance in comparison with somatic allergens.

Diagnostic methods for the study of allergic reactions to Anisakis simplex (A.s.) based on whole-body extracts of the larva are clearly insufficient. To study the allergenicity of the proteins secreted by the parasite. Comparison with somatic antigens and determination of their clinical importance in allergic patients were also addressed. An excretory/secretory (E/S) extract was produced by culturing third-stage A.s. larvae. It was used to perform immediate skin tests and to determine specific IgE in 10 patients diagnosed with allergy to A.s. Both tests were compared with the results obtained with the whole-body extract (somatic (S)). The molecular weight (MW) of their allergens was determined by immunoblotting, and a single-blind placebo-controlled oral challenge with E/S proteins was performed. Finally, allergens' resistance to gastric pepsin and acid pH was explored. A.s. larvae secreted allergens more potent than those present in the S extract. The skin prick test wheal area produced by E/S molecules and the absorbance obtained in the determination of specific IgE with these allergens (ELISA) were 5.8 times bigger than those obtained with S extract. MW allergens of 72 and 56 kDa in E/S extracts and those of 56, 48 and 43 kDa in S extract were recognized by more than 50% of the patients. Partial cross-reactivity between them was revealed by immunoblotting inhibition studies. Oral challenge with E/S extract (up to 479 microg) was negative in all the patients. Treatment of E/S proteins with gastric pepsin inhibited the binding of the E/S allergens for specific IgE. The acid pH did not affect the overall binding of IgE to E/S extract. It decreased by 15.23% and 19.96% at pH 4 and 2, but the difference was not statistically significant. A.s. secretes allergens more potent than somatic antigens and should be used in the diagnostic procedures. These allergens are inactivated by the pepsin, which supports the theory that live larva is necessary to induce an allergic reaction in most of the patients.

Patients with anaphylaxis to pea can have peanut allergy caused by cross-reactive IgE to vicilin (Ara h 1)

Background: Serologic cross-reactivity among legumes has been described; however, it is rarely clinically significant. In this study 3 patients with a history of anaphylaxis to pea are described who subsequently had symptoms after ingestion of peanut. Objective: We investigated whether the peanut-related symptoms were due to cross-reactivity between pea and peanut proteins. Methods: Peanut-related symptoms were documented according to case history or double-blind, placebo-controlled food challenge results. Skin prick tests were performed, and specific IgE levels were determined for pea and peanut with the CAP system FEIA. IgE-binding proteins in pea and peanut were identified by using immunoblot analysis. Cross-reactivity was studied by means of immunoblot and ELISA inhibition studies with whole extracts and purified allergens. Results: Peanut-related symptoms consisted of oral symptoms in all patients, with additional urticaria and dyspnea or angioedema in 2 patients. All patients had a positive skin prick test response and an increased IgE level to pea and peanut. Immunoblotting revealed strong IgE binding to mainly vicilin in pea extract and exclusively to Ara h 1 in crude peanut extract. Immunoblot and ELISA inhibition studies with crude extracts, as well as purified proteins, showed that IgE binding to peanut could be inhibited by pea but not or only partially the other way around. Conclusion: Clinically relevant cross-reactivity between pea and peanut does occur. Vicilin homologues in pea and peanut (Ara h 1) are the molecular basis for this cross-reactivity. (J Allergy Clin Immunol 2003;111:420-4.)