The maize major allergen, which is responsible for food-induced allergic reactions, is a lipid transfer protein.

BACKGROUND: Cereals are the most important nutritional component in the human diet. Food-induced allergic reactions to these substances therefore have serious implications, and exhaustive diagnosis is required. Such diagnosis is still difficult because of the incomplete knowledge about major cereal allergens. In particular, few food-induced allergic reactions to maize have been reported, and no information on the allergenic proteins is available. OBJECTIVE: Having observed several anaphylactic reactions to maize, we planned a study to identify maize major allergens and cross-reactivity with other cereals, as well as to peach because the majority of patients also reacted to Prunoideae fruits. METHODS: Twenty-two patients with systemic symptoms after maize ingestion and positive skin prick test responses and serum-specific IgE antibodies to maize were selected. The IgE-reactivity pattern was identified by SDS-PAGE and immunoblotting. The major allergen identified was then purified by HPLC and characterized by mass spectrometry, determination of the isoelectric point value, and N-terminal amino acid sequencing. RESULTS: Sera from 19 (86%) of the 22 patients recognized a 9-kd protein, thus confirming this as the maize major allergen. This protein had an isoelectric point of greater than 9, a molecular mass of 9047.0 d, and no glycosylation. Determination of its N-terminal sequence showed that it was a lipid transfer protein (LTP). By using immunoblotting-inhibition experiments, we demonstrated that the LTP cross-reacts completely with rice and peach LTPs but not with wheat or barley LTPs. N-terminal sequence of the 16-kd allergen (recognized by 36% of patients) showed it to be the maize inhibitor of trypsin. This protein cross-reacts completely with grass, wheat, barley, and rice trypsin inhibitors. CONCLUSION: The major allergen of maize is an LTP with a molecular weight of 9 kd that is highly homologous with the peach LTP, the major allergen of the Prunoideae subfamily.

Hazelnut allergy: a double-blind, placebo-controlled food challenge multicenter study.

BACKGROUND: Tree nuts are a common cause of food allergy in Europe. However, few studies deal with real food allergy to hazelnuts in subjects believed to be allergic to this food. OBJECTIVE: We sought to select subjects with a history of allergic reactions on ingestion of hazelnut and determine how many of these have true allergy by means of the double-blind, placebo-controlled food challenge (DBPCFC). METHODS: Eighty-six subjects with a history of symptoms after hazelnut ingestion were recruited from 3 allergy centers (Milan, Zurich, and Copenhagen). All subjects underwent skin prick tests (SPTs) with aeroallergens and hazelnut, as well as having their specific hazelnut IgE levels determined. Diagnosis of clinical relevant food allergy was made on the basis of the DBPCFC. RESULTS: Sixty-seven (77.9%) of 86 subjects had a positive DBPCFC result; 8 were placebo responders, and 11 were nonresponders. Of the 11 nonresponders, 4 had positive open-challenge test results. Of the DBPCFC-positive subjects, 87% also had positive skin test responses to birch pollen extract. Specific IgE determination for hazelnut (positive CAP response >/=0.7 kU/L [ie, class 2]) showed a sensitivity of 0.75, a positive predictive value (PPV) of 0.92, a specificity of 0.16, and a negative predictive value (NPV) of 0.05. Skin tests with commercial hazelnut extract produced a sensitivity of 0.89, a PPV of 0.92, a specificity of 0.05, and an NPV of 0.05. Skin tests with natural food produced a sensitivity of 0.88, a PPV of 0.94, a specificity of 0.27, and an NPV of 0.15. CONCLUSION: This study shows that hazelnut is an allergenic source that can cause real food allergy, as confirmed by DBPCFC. Skin and IgE tests demonstrated reasonable sensitivity and PPV but a very low specificity and NPV, thus implying that these should not be used to validate the diagnosis of food allergy to hazelnut.

Evidence for a lipid transfer protein as the major allergen of apricot.

BACKGROUND: Apricots are widely grown in Europe, and allergic reactions are becoming more common, especially oral allergy syndrome. Apricot belongs to the botanical subfamily of Prunoideae, which includes peach, the major allergen of which was identified as a 9-kd protein, a lipid transfer protein (LTP). OBJECTIVE: The aim of the study was to evaluate the IgE reactivity pattern to an apricot extract in subjects with allergic reactions to apricot, as demonstrated by a positive oral challenge response. METHODS: Thirty patients were investigated. All the patients displayed oral allergy syndrome (2 with systemic reactions) to apricot, with positive open food challenge responses, skin prick test responses, and serum-specific IgE antibodies to apricot. The IgE reactivity pattern to apricot extract was identified by using SDS-PAGE and immunoblotting. The major allergen, a 9-kd protein, was then purified by HPLC and characterized by periodic acid-Schiff stain, isoelectric point determination, and N-terminal amino acid sequencing. RESULTS: The sera from all patients allergic to apricot recognized the 9-kd protein, whereas none of the other allergens, with molecular weights from 15 to 80 kd, acted as a major allergen. The 9-kd allergen has an isoelectric point of 8.7 and is not glycosylated. Determination of the N-terminal 34 amino acid sequence showed that it belongs to the LTP family, with a 94% homology with the LTP from peach. IgE blotting of the apricot extract was completely inhibited by the 9-kd purified LTP from peach. CONCLUSIONS: The major allergen of apricot is an LTP, which is highly cross-reactive with the LTP from peach.

Clinical role of a lipid transfer protein that acts as a new apple-specific allergen.

BACKGROUND: Allergy to apple is commonly associated with birch pollinosis because the two share homologous allergens. However, some patients have apple allergy but no birch pollinosis, suggesting that there are allergens that do not cross-react with birch. OBJECTIVE: The aim of the study was to evaluate the IgE reactivity pattern to an apple extract in subjects with allergic reactions to apple, with and without birch hay fever. METHODS: Forty-three patients with oral allergy syndrome for apple and positive open food challenge, skin prick test, and serum specific IgE antibodies to apple were admitted to the study. Thirty-two had birch pollinosis (documented by specific IgE for birch) and 11 were not allergic to birch. The IgE reactivity pattern to apple extract was identified by SDS-PAGE and immunoblotting. The consistent allergen, a 9-kd protein, was then purified by HPLC and characterized by periodic acid-Schiff staining, isoelectric point, and N-terminal amino acid sequencing. RESULTS: The sera from 28% of patients allergic to apple with birch pollinosis, but from all patients allergic only to apple, recognized the 9-kd protein. This protein has an isoelectric point of 7.5 and is not glycosylated. Determination of its partial amino acid sequence showed that it belongs to the family of lipid transfer proteins, which act as major allergens in Prunoideae fruits. CONCLUSIONS: These results indicate that a lipid transfer protein is an important allergen in patients allergic to apple but not to birch pollen. The prevalent IgE reactivity to this allergen in subjects with no birch pollinosis and the physicochemical characteristics of this protein suggest that sensitization may occur through the oral route.

The major allergen of peach (Prunus persica) is a lipid transfer protein.

BACKGROUND: Allergy to fresh fruits and vegetables is mostly observed in subjects with pollinosis, especially from birch, because of cross-reacting allergens in vegetable foods and pollens. However, allergic reactions to fruits, specifically Rosaceae fruits, have been reported in subjects without pollinosis. OBJECTIVE: This study evaluated the pattern of IgE reactivity, identifying the allergen responsible in 2 groups of patients with oral allergy syndrome to peach with or without birch pollinosis. METHODS: The allergenic components of peach were detected by SDS-PAGE and immunoblotting. The major peach allergen was purified by HPLC with a cation-exchange column followed by gel filtration chromatography. Its IgE-binding capacity and its homology with the protein of the crude extract were demonstrated by immunoblotting inhibition techniques. To better characterize this allergen, periodic acid-Schiff stain and isoelectrofocusing were used. The amino acid sequencing was done with a gas-phase sequencer. RESULTS: SDS-PAGE and immunoblotting of the 15 patients allergic to peach, 8 without and 7 with birch pollinosis, showed that they all recognized a protein with a molecular weight of 9 kd. This was the only allergen recognized by patients not sensitized to pollen, whereas the birch pollen-sensitive patients had IgE binding to other allergenic proteins at higher molecular weights. The purified 9-kd protein retained its IgE-binding capacity, was negative to periodic acid-Schiff stain, and had an isoelectric point value of greater than 9. A search in the Swiss Prot Bank showed this was a lipid transfer protein, belonging to a group of molecules involved in the defensive system of plants. CONCLUSIONS: The major allergen of peach is a 9-kd protein belonging to the group of lipid transfer proteins. This is the only allergen recognized by patients allergic to peach but not sensitized to birch pollen.

Sensitization to the major allergen of Brazil nut is correlated with the clinical expression of allergy.

BACKGROUND: Only a few studies have investigated the clinical role of food allergens, especially the relationship between sensitization to a given allergen and occurrence of adverse reactions when eating the relevant food item. OBJECTIVE: This study evaluated the clinical role of the allergens of Brazil nut by comparing the patterns of IgE binding in sera from 11 patients with anaphylaxis after eating Brazil nuts with those from 10 subjects with no symptoms to this food item. Both groups had specific IgE to Brazil nut. METHODS: Allergens in the in-house extract of Brazil nut were identified by SDS-PAGE/immunoblotting, the major allergen was purified by HPLC, and its N-terminal sequence was determined by a protein sequencer. RESULTS: SDS-PAGE/immunoblotting detected a number of allergenic components with molecular weights ranging from 4 to 58 kd. All sera from symptomatic patients recognized a 9-kd allergen corresponding (as established by amino acid sequencing) to a 2S albumin already described as a major allergen of Brazil nut, whereas the other allergens each bound IgE from less than 50% of sera. No sera from asymptomatic subjects showed IgE binding to the 9-kd allergen, but they did recognize components from 25 to 58 kd, which are minor allergens. CONCLUSIONS: These findings indicate that the allergen underlying clinical reactions to Brazil nut is a 2S albumin of 9 kd and that in vitro reactivity to this allergen identifies subjects who react in vivo to ingestion of this food.

Identification of actinidin as the major allergen of kiwi fruit.

BACKGROUND: Allergic reactions to fruits and vegetables are among the most frequent food allergies in adults. Kiwi fruit (Actinidia chinensis) is commonly involved, causing local mucosal, systemic, or both types of symptoms by an IgE-mediated mechanism. In a previous study on 30 patients allergic to kiwi, we identified a major allergen of 30 kd against which all sera tested clearly reacted. Other allergens were detected at 12, 24, and 28 kd. OBJECTIVE: The aim of this study was to fully characterize the major kiwi fruit allergen of 30 kd. METHODS: Allergens were separated and purified by high-performance liquid chromatography with anion-exchange columns. The purity of the single proteins was checked by sodium dodecylsulfate-polyacrylamide gel electrophoresis, and their allergenicity was checked by immunoblotting with a pool of sera from patients allergic to kiwi. The allergens were characterized by isoelectrofocusing and amino acid sequencing, and periodic acid-Schiff stain was used to detect glycoproteins. RESULTS: Proteins of 30, 28, 24, and 17 kd were purified by high-performance liquid chromatography. IgE binding indicated the 30 kd protein, which showed an isoelectric point of 3.5, as the major allergen of kiwi. Determination of its partial amino acid sequence and comparison with the Swiss Protein Bank showed that this was actinidin, the main protein component of kiwi. The 24 and 28 kd proteins had the same N-terminal sequence, which did not correspond to any known protein. The 17 kd protein had a blocked N-terminal sequence. CONCLUSIONS: These results demonstrate that the major allergen of kiwi fruit, Act c 1, is actinidin, a proteolytic enzyme belonging to the class of thiol-proteases. Two other allergens of 24 and 28 kd appear identical on amino acid sequencing.

Identification of the allergenic components of kiwi fruit and evaluation of their cross-reactivity with timothy and birch pollens.

BACKGROUND: Only a few food allergens have as yet been identified, mainly because of the difficulty of obtaining a sufficient number of patients who are clinically sensitized to a given food. This is more feasible in the case of the oral allergy syndrome (OAS), a common form of food allergy, which is especially prevalent in patients with pollinosis. OBJECTIVE: We designed a study to identify the allergens of kiwi fruit (Actinidia chinensis) by analyzing the sera of patients with OAS for kiwi and to examine the cross-reactivity of these allergens with timothy and birch pollen allergens. METHODS: Twenty-seven patients with OAS for kiwi, a positive skin prick test response and serum IgE antibody to kiwi, and a positive open kiwi challenge test result and three patients who had OAS with severe systemic symptoms, which excluded a challenge test, were included in this study. The different polypeptide components of an extract of fresh kiwi were separated by sodium dodecylsulfate-polyacrylamide gel electrophoresis and analyzed by IgE immunoblotting with sera from these patients. Cross-reactivity with the two pollen extracts was assessed by inhibition of the immunoblots with pooled and individual patients' sera. RESULTS: Twelve IgE-binding components with molecular weights ranging from 12 to 64 kd were identified in the kiwi extract, but only a 30 kd component acted as major allergen, being recognized by sera of 100% of these patients. Inhibition of kiwi immunoblots with timothy and birch pollen extracts demonstrated strong cross-reactivity with some of the kiwi allergens, suggesting complete identity between certain food and pollen allergens; whereas others, particularly the 30 kd allergen, were only partially inhibited, suggesting much weaker cross-reactivity. CONCLUSIONS: Kiwi fruit contains a large number of allergens widely cross-reacting with allergens in grass and birch pollen extracts. Nevertheless, the major allergen at 30 kd appears to be specific for kiwi.

Comparison of results of skin prick tests (with fresh foods and commercial food extracts) and RAST in 100 patients with oral allergy syndrome.

One hundred adult patients with a history of oral allergy syndrome (OAS) after ingestion of fruits and vegetables, 77 patients with hay fever and 13 with skin prick tests and RAST positive to pollens but without seasonal symptoms, and 32 normal nonallergic control subjects, had Phadebas RAST and skin prick tests with commercial extracts (CSPT) and with fresh foods (FFSPT) to assess the reliability of these three tests. Sensitivity was better with FFSPT for carrot, celery, cherry, apple, tomato, orange, and peach; better with CSPT for peanut, pea, and walnut; and better with RAST for hazelnut. Specificity, negative predictive value, and positive predictive value of the three tests were determined for apple, carrot, hazelnut, orange, pea, peanut, and tomato. Specificity in the patient groups ranged between 40% (pea) and 100% (apple) for CSPT, between 61% (peanut) and 87% (carrot) for RAST, and between 42% (carrot) and 93% (peanut) for FFSPT. However, all tests were negative in the control group. Thus, false positive results may result from cross-reactivity with pollen allergens. The diagnostic accuracy of these tests in the population with OAS proved comparable for peanut, carrot, hazelnut, and pea. FFSPT proved more sensitive than CSPT or RAST in confirming a history of OAS to certain alimentary allergens, such as apple, orange, tomato, carrot, cherry, celery, and peach.

The oral allergy syndrome.

We have studied 262 patients suffering from hay fever and oral allergy syndrome after fruit and vegetable ingestion. The history and the results of RAST and skin test for pollen showed a contemporary presence of systemic and local symptoms in most of the patients; a close connection between age of onset of hay fever and oral allergy syndrome; a frequent association between allergy to some pollens and some vegetables, such as between apple, carrot, pear, cherry with birch pollen and tomato, melon, watermelon with grass pollen.

Food allergy diagnosis protocol.

The protocol is proposed in a definite way for recurrent or chronic symptoms. In the first case, the diagnosis is made by PRICK, RAST and provocation tests with suspect foods. On 100 patients, the reliability of PRICK and RAST has been shown. In the second case, a non-allergenic diet and the recurrence of symptoms on the re-introduction of certain foods has permitted isolation of the causative foods and the differentiation of food allergy and intolerance. This has been done with 386 cases of urticaria and angio-oedema, where it has been shown that 1.9% of the reactions had a food-linked origin.

IgE-mediated food allergy.

The behaviour of some immunologic parameters was studied in order to explain the presence or the lack of symptomatology in subjects with IgE specific to foods. In particular, we compared the levels of serum and salivary IgA and IgE antibody specific to apple in 33 patients allergic to birch and with anti-apple IgE, of whom 24 had oral allergy syndrome after eating apple and nine were apple tolerants. We found no differences in the IgE-related parameters such as levels of anti-apple IgE antibody in serum and secretions, mean size of apple skin prick test, and number of patients with a positive histamine release from peripheral basophils stimulated by apple. Similarly, we detected no statistically significant differences in serum and salivary total IgG and total anti-apple IgA antibody of the two groups. We could, however, observe a significantly higher anti-apple salivary IgA/anti-apple serum IgE and anti-apple salivary IgA/anti-apple serum IgA ratios and a significant correlation between the same parameters in tolerant patients when compared with the intolerants. These results seem to suggest that the presence of appropriate levels of secretory anti-apple IgA associated to anti-apple IgE might be one of the factors playing a key role in the prevention of clinical symptoms in atopic subjects.