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Routine diagnostic methods for allergies to plant-derived foods are based on skin prick test (SPT) with commercial extracts, prick-by-prick (PbP) with fresh food, serum-specific IgE measurement, and oral food challenge.We discuss the possibility and the advantages of performing, in patients with oral allergy syndrome (OAS) by fruit and vegetables (excluding nuts) PR-10 allergy, component-resolved diagnosis (CRD) by SPT and PbP with raw and cooked vegetables, rather than performing a CRD with in vitro tests by drawing blood.Based on our clinical experience and the studies published in the literature, we believe that, at least for the OAS by fruit and vegetables (excluding nuts) PR-10 allergy, the search for sensitizing allergens and related cross-reactive allergens with SPT and PbP can be performed routinely in clinical practice, even at the primary-care level., Competing Interests: None

Plant lipid transfer proteins (LTPs) are widespread plant food allergens, highly resistant to food processing and to the gastrointestinal environment, which have been described as the most common food allergens in the Mediterranean area. LTP allergy is widely described in adults, but it represents an emerging allergen also in the pediatric population. Little is known about the real prevalence and the clinical features of this allergy in children and it still often remains underdiagnosed in these patients. An early identification and a deeper knowledge of this allergy in childhood can avoid severe systemic reactions and improve the child's quality of life. Pediatricians should always consider the possibility of LTP involvement in cases of plant-derived food allergy., (Copyright © 2020 SEICAP. Published by Elsevier España, S.L.U. All rights reserved.)

Scope: Chickpea (Cicer arietinum) allergy has frequently been reported particularly in Spain and India. Nevertheless, chickpea allergens are poorly characterized. The authors aim to identify and characterize potential allergens from chickpea., Methods and Results: Candidate proteins are selected by an in silico approach or immunoglobuline E (IgE)-testing. Potential allergens are prepared as recombinant or natural proteins and characterized for structural integrity by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), circular dichroism (CD)-spectroscopy, and mass spectrometry (MS) analysis. IgE-sensitization pattern of Spanish chickpea allergic and German peanut and birch pollen sensitized patients are investigated using chickpea extracts and purified proteins. Chickpea allergic patients show individual and heterogeneous IgE-sensitization profiles with extracts from raw and boiled chickpeas. Chickpea proteins pathogenesis related protein family 10 (PR-10), a late embryogenesis abundant protein (LEA/DC-8), and a vicilin-containing fraction, but not 2S albumin, shows IgE reactivity with sera from chickpea, birch pollen, and peanut sensitized patients. Remarkably, allergenic vicilin, DC-8, and PR-10 are detected in the extract of boiled chickpeas., Conclusion: Several IgE-reactive chickpea allergens are identified. For the first time a yet not classified DC-8 protein is characterized as minor allergen (Cic a 1). Finally, the data suggest a potential risk for peanut allergic patients by IgE cross-reactivity with homologous chickpea proteins., (© 2020 Wiley-VCH GmbH.)

Purpose of Review: The route of allergen sensing via the skin appears to influence the immune system towards mounting a type 2 response, especially in genetically predisposed individuals. Allergens recognized this way may derive from microbial, animal, food, or other plant sources and trigger atopic dermatitis. Allergens can be grouped into families depending on their structure and function, harboring significant structural and sequence similarities. Cross-reactivity between allergens is believed to arise as a consequence, and to underlie the development of further atopic diseases., Recent Findings: Especially for the plant allergens of the families of PR10-related proteins and profilins, immune cross-reactions have been described. Actual studies support that food and pollen allergens can aggravate skin lesions in patients suffering from atopic dermatitis. Further on, allergens derived from air-borne or skin-borne fungi belong to common allergen families and bear cross-reactivity potential. Cross-reactivity to human homologous proteins, so-called autoallergens, is discussed to contribute to the chronification of atopic dermatitis., Summary: Due to high evolutionary conservation, allergic reactions can be triggered by highly homologous members of allergen families on the humoral as well as on the cellular level.

Background & Aims: Wheat has become the world's major staple and its consumption correlates with prevalence of noncommunicable disorders such as inflammatory bowel diseases. Amylase trypsin inhibitors (ATIs), a component of wheat, activate the intestine's innate immune response via toll-like receptor 4 (TLR4). We investigated the effects of wheat and ATIs on severity of colitis and fecal microbiota in mice., Methods: C57BL/6 wild-type and Tlr4 -/- mice were fed wheat- or ATI-containing diets or a wheat-free (control) diet and then given dextran sodium sulfate to induce colitis; we also studied Il10 -/- mice, which develop spontaneous colitis. Changes in fecal bacteria were assessed by taxa-specific quantitative polymerase chain reaction and 16S ribosomal RNA metagenomic sequencing. Feces were collected from mice on wheat-containing, ATI-containing, control diets and transplanted to intestines of mice with and without colitis on control or on ATI-containing diets. Intestinal tissues were collected and analyzed by histology, immunohistochemistry, and flow cytometry. Bacteria with reported immunomodulatory effects were incubated with ATIs and analyzed in radial diffusion assays., Results: The wheat- or ATI-containing diets equally increased inflammation in intestinal tissues of C57BL/6 mice with colitis, compared with mice on control diets. The ATI-containing diet promoted expansion of taxa associated with development of colitis comparable to the wheat-containing diet. ATIs inhibited proliferation of specific human commensal bacteria in radial diffusion assays. Transplantation of microbiota from feces of mice fed the wheat- or ATI-containing diets to intestines of mice on control diets increased the severity of colitis in these mice. The ATI-containing diet did not increase the severity of colitis in Tlr4 -/- mice., Conclusions: Consumption of wheat or wheat ATIs increases intestinal inflammation in mice with colitis, via TLR4, and alters their fecal microbiota. Wheat-based, ATI-containing diets therefore activate TLR4 signaling and promote intestinal dysbiosis., (Copyright © 2020 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Background & Aims: Pro-inflammatory biomarkers are well-established contributors to insulin resistance and represent valid targets for diabetes management and prevention. Yet, little is known whether nutrition could play a role in modulating various aspects of immune-inflammatory responses. Our aim is to assess the effect of isocaloric animal and plant protein dietary interventions on selected biomarkers representing various immune-inflammatory pathways., Methods: We enrolled 37 participants with type 2 diabetes (age 64 ± 6 years, body mass index 30.2 ± 3.6 kg/m 2 , glycated hemoglobin 7.0 ± 0.6%) who underwent an either high-animal protein (AP) or high-plant protein (PP) diet (30 E% protein, 40 E% carbohydrates, 30 E% fat) for 6-weeks. Clinical examinations were performed at beginning and end of the study. Levels of pro-inflammatory adipokines [chemerin, progranulin], cytokines [tumor necrosis factor α (TNF-α), interleukin 6 (IL-6), soluble urokinase-type plasminogen activator receptor (suPAR), transforming growth factor beta 1 (TGF-β1)], and proteins [calprotectin, lactoferrin and growth differentiation factor 15 (GDF-15)] were determined in blood serum using enzyme-linked immunosorbent assay., Results: Chemerin and progranulin concentrations decreased following AP and PP diets. TGF-β1 increased in AP and decreased in PP, whereas calprotectin increased in PP and decreased in AP. No statistically significant differences in the concentrations of IL-6, TNF-α, suPAR, lactoferrin and GDF-15 could be seen in either of the protein diet arms., Conclusions: These results suggest that both AP and PP diets may effectively reduce the levels of the pro-inflammatory adipokines chemerin and progranulin. The effects on the additional immune-inflammatory biomarkers seem to be more complex., Clinical Trial Registry Number: NCT02402985 (ww.clinicaltrials.gov)., (Copyright © 2019. Published by Elsevier Ltd.)

An 8-week growth trial was conducted to investigate the effects of replacing dietary fishmeal with a plant protein blend on the growth performance, mucosal barrier integrity and the related regulation mechanism in Amur Sturgeon (Acipenser schrenckii) with initial weight of 87.48 g. Three isonitrogenous and isoenergetic diets were prepared. A basal diet containing 540 g/kg fishmeal (P0), whereas the other two diets were formulated by replacing 50% and 100% of FM with plant protein blend (soybean protein concentrate and cottonseed protein concentrate), and named as P50 and P100, respectively. Although essential amino acids, fatty acids, and available phosphorus had been balanced according to the nutrient requirement of sturgeon, compared with the fish of P0 and P50, the full plant protein diet (P100) significantly reduced growth performance and survival, and accompanied with serious spiral valve intestinal (SVI) damage. The increased tissue necrosis and failed responses in anti-oxidation, programming apoptosis, autophagy and cell proliferation system were regulated by inhibiting ERK1 phosphorylation, which indicated that SVI hypoimmunity and functional degradation were the main reasons for the high mortality and low utilization ability of plant protein in Amur sturgeon., (Copyright © 2019. Published by Elsevier Ltd.)

The wheat varietal selection undertaken by breeders in recent decades has been tailored mainly to improve technological and productivity-related traits; however, the latter has resulted in a considerable impoverishment of the genetic diversity of wheat-based products available on the market. This pitfall has encouraged researchers to revalue the natural diversity of cultivated and non-cultivated wheat genotypes in light of their different toxic/immunogenic potential for celiac disease and wheat-allergic patients. In the present investigation, an advanced proteomic approach was designed for the global characterization of the protein profile of selected tetraploid wheat genotypes ( Triticum turgidum ). The approach combined proteins/peptides sequence information retrieved by specific enzymatic digestions (single and dual proteolytic enzymes) with protein digestibility information disclosed by means of in-vitro simulated human gastroduodenal digestion experiments. In both cases, the peptide pools were characterized by discovery analysis with liquid chromatography high-resolution tandem mass spectrometry, and specific amino acid sequences were identified via commercial software. The peptide list was screened for in silico toxicity/immunogenicity risk assessment, with the aid of various open-source bioinformatics tools for epitopes matching. Given the global information provided by the designed proteomic approach, the in silico risk assessment not only tackled toxicity implication for celiac disease patients, but also scouted for immunogenic sequences relevant for wheat allergic patients, achieving a comprehensive characterization of the protein profile of the selected genotypes. These latter were assessed to encrypt a variable number of toxic/immunogenic epitopes for celiac disease and wheat allergy, and as such they could represent convenient bases for breeding practices and for the development of new detoxification strategies.

Summary: Dietary lectins play a major role in the activation of mast cells / basophils by bridging cell surface IgE glycans to release histamine and other mediators. In the present study, the effect of mannose / glucose-specific banana lectin (BanLec) on the activation of mast cells / basophils from non-atopic and atopic subjects has been investigated. BanLec was purified from banana pulp in a yield of 7 mg/kg. Leukocytes isolated from heparinized blood of non-atopic / atopic subjects were used for quantitation of the released histamine. Approximately 28.2% of the atopics (n = 117) was positive by skin prick test (SPT) to purified BanLec (100 µg/mL concentration), and all the non-atopics (n = 20) were negative. Maximal release of histamine was seen at 2 µg of BanLec. In percent histamine release, an increase of 35-40% is observed in case of atopics (n = 7) compared to non-atopics (n = 5), and the histamine release from atopic and non-atopic subjects correlates fairly well with the total serum IgE levels (R2 = 0.817). BanLec also induces release of histamine (26.7%) from mast cells present in rat peritoneal exudate cells. BanLec can significantly activate and degranulate mast cells and basophils by cross-linking the trimannosidic core mannose of IgE glycans in atopic population as compared to non-atopic population; the activation is marginal in the case of non-atopics.

Cow's milk protein allergy (CMPA) is the most common food allergy in infancy. Non-IgE mediated (NIM) forms are little studied and the responsible mechanisms of tolerance acquisition remain obscure. Our aim was to study the intestinal microbiota and related parameters in the fecal samples of infants with NIM-CMPA, to establish potential links between type of formula substitutes, microbiota, and desensitization. Seventeen infants between one and two years old, diagnosed with NIM-CMPA, were recruited. They were all on an exclusion diet for six months, consuming different therapeutic protein hydrolysates. After this period, stool samples were obtained and tolerance development was evaluated by oral challenges. A control group of 10 age-matched healthy infants on an unrestricted diet were included in the study. Microbiota composition, short-chain fatty acids, calprotectin, and transforming growth factor (TGF)-β₁ levels were determined in fecal samples from both groups. Infants with NIM-CMPA that consumed vegetable protein-based formulas presented microbiota colonization patterns different from those fed with an extensively hydrolyzed formula. Differences in microbiota composition and fecal parameters between NIM-CMPA and healthy infants were observed. Non-allergic infants showed a significantly higher proportion of Bacteroides compared to infants with NIM-CMPA. The type of protein hydrolysate was found to determine gut microbiota colonization and influence food allergy resolution in NIM-CMPA cases.

Despite a huge number of studies, many aspects of the lipid transfer protein (LTP) syndrome, the most frequent primary food allergy in Mediterranean countries, remain unclear. Its peculiar geographical distribution, along with the extreme variability of its clinical expression, makes this type of food allergy something unique in the panorama of IgE-mediated food-induced allergic reactions. This review article tried to summarize the current knowledge about the most important aspects of LTP sensitization and allergy, along with the importance of positive and negative co-factors in the clinical expression of the syndrome as well as the issues regarding the cross-reactivity between LTPs present in botanically related and unrelated foods. Further, the possible absence of the protein from some plant foods is discussed., (© 2017 John Wiley & Sons Ltd.)

Inclusion of prebiotics in aqua feeds, though a costly strategy, has increased as a means to improve growth. Still, its effects on health improvement are not fully disclosed. Regarding their immunestimulatory properties, research has focused on carbohydrates such as fructooligosaccharides and xylooligosaccharides demonstrating their modulatory effects on immune defences in higher vertebrates but few studies have been done on their impact on fish immunity. Replacing fish meal (FM) by plant protein (PP) sources is a current practice in the aquaculture business but their content in antinutrients is still a drawback in terms of gut well-functioning. This work intends to evaluate the short-term effect (7 or 15 days feeding the experimental diets) on juvenile European seabass (Dicentrarchus labrax) immune status of dietary i) replacement of FM by PP sources; ii) prebiotics supplementation. Six isoproteic (46%) and isolipidic (15%) diets were tested including a FM control diet (FMCTRL), a PP control diet (PPCTRL, 30 FM:70 PP) and four other diets based on either FM or PP to which short-chain fructooligosaccharides (scFOS) or xylooligosaccharides (XOS) were added at 1% (FMFOS, PPFOS, FMXOS, PPXOS). The replacement of FM by PP in the diets induced nitric oxide (NO) and lysozyme production, while immunoglobulins (Ig), monocytes percentage and gut interleukin 10 (IL10) gene expression were inhibited. Dietary scFOS supplementation inhibited total bactericidal activity and neutrophils relative percentage regardless protein source and increased plasma NO and thrombocytes percentage in fish fed FM-based diets, while monocytes percentage was increased in PPFOS-fed fish. XOS supplementation down-regulated immune gene expression in the gut while it partly enhanced systemic response. Inconsistency among results regarding FM replacement by PP-based ingredients exposes the need for further research considering both local and systemic responses. Distinct outcomes of prebiotic supplementation were highlighted reflecting sight-specific effects with no clear interaction with protein source., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

To solve the future food insecurity problem, alternative and sustainable protein sources (e.g. insects, rapeseed, fava bean and algae) are now being explored for the production of food and feed. To approve these novel protein sources for future food a comprehensive risk assessment is needed according to the European food legislation. Allergenicity risk assessment might pose some major difficulties, since detailed guidance on how to assess the allergenic potential of novel foods is not available. At present, the approach relies mostly on the guidance of allergenicity assessment for genetically modified (GM) plant foods. The most recent one was proposed by EFSA (2010 and 2011); "weight-of-evidence approach". However this guidance is difficult to interpret, not completely applicable or validated for novel foods and therefore needs some adjustments. In this paper we propose a conceptual strategy which is based on the "weight-of-evidence approach" for food derived from GM plants and other strategies that were previously published in the literature. This strategy will give more guidance on how to assess the allergenicity of novel food proteins and protein sources., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Plant allergens, being one of the most widespread allergenic substances, are hard to avoid. Hence, their identification and characterization are of prime importance for the diagnosis and treatment of food allergy. The reported allergies to fruits mainly evoke oral allergy syndrome caused by the presence of cross-reactive IgE to certain pollens and thus, allergy to fruits has also been linked to particular pollens. Many fruit allergies are being studied for their causative allergens, and are being characterized. Some tropical or exotic fruits are responsible for region-specific allergies for which only limited information is available, and generally lack allergen characterization. From a survey of the literature on fruit allergy, it is clear that some common fruits (apple, peach, musk melon, kiwi fruit, cherry, grape, strawberry, banana, custard apple, mango and pomegranate) and their allergens appear to be at the center of current research on food allergy. The present review focuses on common fruits reported as allergenic and their identified allergens; a brief description of allergens from six rare/tropical fruits is also covered.

Background: We previously reported the initial results of the first multicenter, randomized, double-blind, placebo-controlled clinical trial of peanut sublingual immunotherapy (SLIT), observing a favorable safety profile associated with modest clinical and immunologic effects in the first year., Objective: We sought to provide long-term (3-year) clinical and immunologic outcomes for our peanut SLIT trial. Key end points were (1) percentage of responders at 2 years (ie, could consume 5 g of peanut powder or a 10-fold increase from baseline), (2) percentage reaching desensitization at 3 years, (3) percentage attaining sustained unresponsiveness after 3 years, (4) immunologic end points, and (5) assessment of safety parameters., Methods: Response to treatment was evaluated in 40 subjects aged 12 to 40 years by performing a 10-g peanut powder oral food challenge after 2 and 3 years of daily peanut SLIT therapy. At 3 years, SLIT was discontinued for 8 weeks, followed by another 10-g oral food challenge and an open feeding of peanut butter to assess sustained unresponsiveness., Results: Approximately 98% of the 18,165 doses were tolerated without adverse reactions beyond the oropharynx, with no severe symptoms or uses of epinephrine. A high rate (>50%) discontinued therapy. By study's end, 4 (10.8%) of 37 SLIT-treated participants were fully desensitized to 10 g of peanut powder, and all 4 achieved sustained unresponsiveness. Responders at 2 years showed a significant decrease in peanut-specific basophil activation and skin prick test titration compared with nonresponders., Conclusions: Peanut SLIT induced a modest level of desensitization, decreased immunologic activity over 3 years in responders, and had an excellent long-term safety profile. However, most patients discontinued therapy by the end of year 3, and only 10.8% of subjects achieved sustained unresponsiveness., (Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Objective: To investigate the reactivity of monoclonal anti-citrullinated protein antibody (ACPA) obtained from peripheral blood B cells of rheumatoid arthritis (RA) patients with human autoantigens as well as environmental proteins by determining the essential epitope for the ACPA., Methods: A human monoclonal ACPA (cyclic citrullinated peptide antibody 1 [CCP-Ab1]) was obtained by screening peripheral blood lymphocytes from 31 patients with RA using a novel monoclonal antibody-secreting cell (ASC) screening system, the immunospot-array assay on a chip. The essential epitope for CCP-Ab1 was determined using epitope mapping. Then, human, microbial, and plant proteins that share the essential epitope identified were searched using BLAST. Finally, representative proteins identified by the search were produced in vitro, and their reactivity with CCP-Ab1 was examined., Results: CCP-Ab1 bound CCP in a citrulline-indispensable manner. In CCP, the 6 amino acid residues required for CCP-Ab1 binding were identified. In the BLAST search, 38 human, 56 viral, 1,383 fungal, 547 bacterial, and 1,072 plant proteins were found to share the essential epitope, and CCP-Ab1 reacted with all of the recombinant citrullinated proteins tested, which included the various environmental factors, such as various plant proteins that are part of the daily diet., Conclusion: Our findings demonstrate, for the first time, that a monoclonal ACPA (CCP-Ab1) derived from RA patients cross-reacts not only with various autoantigens but also with numerous plant and microbial proteins. We propose that countless environmental factors, including microbes and diet, may trigger the generation of ACPAs that then cross-react with various citrullinated human autoantigens through molecular mimicry to induce RA., (© 2015, American College of Rheumatology.)

Background: Recent studies have highlighted the importance of extra-intestinal routes of sensitization to food-related allergens as the cause of epidemics of food allergy. Instances of Japanese women developing food allergy to wheat after exposure to hydrolyzed wheat protein (HWP) present in facial soap have been reported. However, the epidemiologic impact of these ingredients as a cause of food allergy has not been well studied., Methods: To clarify the epidemiological relationship between food allergy to wheat and contact exposure to HWP, a case-control study of Japanese women aged 20-54 years with self-reported wheat allergy (WA) (cases, n = 157) and age-matched control subjects without WA (controls, n = 449) was performed using a large-scale Web-based research panel. Subjects answered a Web-based questionnaire regarding the use of skin and hair care products, as well as other possible risk factors., Results: Current use of an HWP-containing facial soap (Cha no Shizuku; Yuka) was significantly associated with an increased risk of WA (adjusted odds ratio, 2.6; 95% confidence interval, 1.2-5.7; frequencies of current use in cases and controls; 11% and 6%, respectively). Use of Cha no Shizuku was more common in subjects with more recent-onset WA, implying that this soap may have contributed to the recent epidemic of WA., Conclusions: An epidemiological relationship between WA and contact exposure to HWP has been documented. This study implicates a possible role of contact exposure to food-derived protein hydrolysates as a risk factor for the development of food allergy manifesting itself as anaphylaxis., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Replacement of peanut extracts by recombinant peanut components is an important step in allergy serologic testing. Criteria are needed for the unbiased inclusion of patients into a study to validate such a replacement., Methods: Plasma samples from 64 peanut-positive children (42 reactors, 22 nonreactors in a double-blind, placebo-controlled food challenge) were used to compare IgE reactivity to six recombinant peanut allergens with reactivity to natural peanut proteins extracted at neutral or low pH. We tested the hypothesis that poor extractability of Ara h 9 and other basic allergens at neutral pH leads to under-representation of patients with such sensitization., Results: IgE reactivity to the components did not fully explain IgE reactivity to peanut extract in 5 of 32 reactors with IgE to peanut extract ≤100 kUA /l. IgE reactivity to components was stronger than to the extract in 11 plasma samples, which was largely due to a low Ara h 8 reactivity of the extract. IgE reactivity to Ara h 9 was much lower than reactivity to other basic proteins, some of which bound IgE well in the RAST, but lost IgE reactivity upon immunoblotting., Conclusions: Conventional peanut extracts are deficient in significant IgE-binding components. The inclusion of patients for a validation study should be based on serology performed with improved peanut reagents to avoid a bias against these under-represented, potentially important allergens. To judge clinical relevance of an allergen, the reagent used for inclusion of patients needs to be efficient in detecting IgE to this component., (© 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Seed storage proteins (SSP; Ara h 1, Ara h 2, Ara h 3) have been shown to be major peanut allergens, although recently, peanut lipid transfer protein has been reported to be an important allergen in the Mediterranean area. We sought to investigate the sensitization pattern to peanut SSP and vegetable pan-allergens in a group of peanut-allergic children compared with a peanut-tolerant group., Methods: One hundred and twenty-three children who presented with food allergy were included in the study. Tolerance to peanut ingestion was assessed. Specific IgE was determined by ImmunoCAP, and microarray ISAC was performed. Sensitization frequencies and levels of specific IgE were compared between groups., Results: Fifty-five of 123 children presented symptoms upon contact or ingestion. Frequency of sensitization to Ara h 1, Ara h 2, and Ara h 3 was 60.0%, 72.7%, and 43.6%, respectively, in the group of allergic children vs. 7.4%, 1.5%, and 7.4% in the group of tolerant children. Levels of specific IgE against Ara h 1, Ara h 2, and Ara h 3 were significantly higher in the allergic group (p < 0.001). The frequency of sensitization and the levels of specific IgE against Cor a 8 (36.4% vs. 16.2%) were significantly higher in the allergic children, whereas no significant differences were found for Pru p 3. No differences were seen for other pan-allergens. Patients sensitized to SSP, regardless of sensitization to nsLTP, were allergic rather than tolerant., Conclusion: In our population, peanut-allergic children were mainly sensitive to SSP. A few patients were also sensitive to some nsLTPs. No differences were shown in other pan-allergens., (© 2012 John Wiley & Sons A/S. Published by Blackwell Publishing Ltd.)

Garlic is used all over the world for treatment of different diseases. A wide range of biological activities of garlic has been verified in vitro and in vivo. One of major proteins of garlic which has been isolated and purified is the 14 kDa protein. This protein has been shown to have immunomodulatory effects. In this study, the effect of the 14 kDa protein isolated from aged garlic extract (AGE) was investigated on maturation and immunomodulatory activity of dendritic cells (DC). Proteins were purified from AGE by biochemical method; the semi-purified 14 kDa protein was run on gel filtration Sephadex G50 and its purity was checked by SDS-PAGE. DC were isolated from spleen of BALB/c mice by Nycodenz centrifugation and their adhesiveness to plastic dish. 14 kDa protein from AGE was added to overnight culture of DC medium and the expression percentage of CD40, CD86, and MHC-II was evaluated by flowcytometric analysis. Also, proliferation of T-cells was measured by allogenic mixed lymphocyte reaction (MLR) test. The purified 14 kDa protein isolated from AGE increased the expression of CD40 molecule on DC, but it did not influence CD86 and MHCII molecules. Furthermore, no significant differences were noticed in the pulsed-DC with 14 kDa protein and non-pulsed DC on the MLR., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Background: It is not known why some foods sensitizing via the gastrointestinal tract are prevalent allergenic foods and others are not. Eating habits, processing, and the food matrix have been suggested to influence the allergenicity of a given food. Factors related to protein structure, such as stability to digestion, have also been suggested. 7S globulins from peanut, hazelnut, soy, and pea were studied to determine whether related proteins would induce a similar sensitization when removed from their 'normal' matrix., Methods: Brown Norway rats (soy tolerant or nontolerant) were immunized i.p. 3 times with 100 μg purified peanut, hazelnut, soy, or pea 7S without adjuvant. Sera were analyzed for specific antibodies by different ELISAs (IgG1, IgG2a, and IgE), inhibition ELISA, and rat basophilic leukemia cell assay., Results: The 4 related 7S globulins induced a response with an almost identical level of specific antibodies, but peanut 7S induced IgE of higher avidity than hazelnut and pea 7S which, again, had a higher avidity than IgE induced by soy 7S. Soy tolerance reduced the functionality of IgE without influencing antibody titers., Conclusions: Although the 4 7S globulins are structurally related allergens, they induce antibodies with different antigen-binding characteristics. Peanut 7S induces IgE of a higher avidity than hazelnut and pea 7S which, again, has a higher avidity than IgE induced by soy 7S. We also show that soy tolerance influences the function of antibodies to peanut 7S. These findings may help explain how antibodies of different clinical significances can develop in different individuals sensitized to the same allergen., (Copyright © 2011 S. Karger AG, Basel.)

Background: Tree nut allergy is one of the common potentially life-threatening food allergies in children and adults. Recombinant food allergens offer new perspectives to solve problems of clinical and molecular allergology in diagnosis, research, and therapy of food allergies. So far, superoxide dismutase (s) has been identified as a panallergen and studied in different allergenic sources. Manganese Superoxide Dismutase (MnSOD) has also been reported in pistachio that may cause allergic reactions in atopic subjects. The aim of this study was to describe the cloning, expression, and purification of MnSOD from pistachio nut., Methods: The pistachio MnSOD was cloned and expressed in E. coli BL21 (DE3) using a vector pET-32b (+). A recombinant protein was purified by metal precipitation. The protein immunoreactivity was evaluated using patients' IgE binding by means of ELISA and immunoblotting assays., Results: The MnSOD gene from pistachio was successfully cloned and expressed in E. coli. The purified pistachio MnSOD was recognized by IgE in 10 (40%) out of the 25 sera tested. Our results also showed that this protein might trigger some cross-reactions toward IgE antibodies and thus could be considered as a panallergen., Conclusions: For the first time recombinant manganese superoxide dismutase from nut source was expressed as a possible allergen. This pistachio allergen could be a possible basis for cross-reactivity with MnSOD from other sources.

The present review summarizes current knowledge about lupine allergy, potential sensitization routes, cross-reactions between lupine and other legumes, and the respective IgE-binding proteins. Since the 1990 s, lupine flour is used as a substitute for or additive to other flours, mostly wheat flour, in several countries of the EU. In 1994, the first case of an immediate-type allergy after ingestion of lupine flour-containing pasta was reported. Since then, the number of published incidents following ingestion or inhalation of lupine flour is rising. So far, the Lupinus angustifolius beta-conglutin has been designated as the allergen Lup an 1 by the International Union of Immunological Societies Allergen Nomenclature Subcommittee. Initially, publications focussed on the fact that peanut-allergic patients were at risk to develop anaphylaxis to lupine due to cross-reactivity between peanut and lupine. At present, however, the ratio between cases of pre-existing legume allergy (mostly peanut allergy) to de novo sensitization to lupine seed is nearly 1:1. Although in December 2006, lupine and products thereof were included in the EU foodstuff allergen list according to the Commission Directive 2006/142/EC amending Annex IIIA of Directive 2000/13/EC in order to prevent severe reactions caused by "hidden food allergens", the majority of patients and medical personnel are still not aware of raw lupine seed as potentially dangerous food allergen.

Analytical methods are needed for measuring the levels of protein from allergenic food transferred into cooking oil. A simple method for determination of total protein in cooking oils was developed. Oil was extracted with phosphate-buffered saline with 0.05% Tween (PBST) and the extracts were partitioned with hexane to remove residual oil. Total protein in the PBST extracts was assayed with bicinchoninic acid (BCA), micro-BCA, reducing-agent compatible BCA and CB-XT kits. These methods were used to measure recovery of protein from peanut butter spikes of soy and peanut oil in the range of 50-1000 ppm. Recoveries were generally above 70%. However, the BCA and micro-BCA assays were subject to interference and enhanced color formation which were probably due to co-extracted antioxidants present in oil. The reducing agent-compatible BCA and CB-X protein assays reduced interference and gave lower protein values in crude, cold-pressed, and refined peanut oils. Heating oil to 180 degrees C before extraction also reduced interference-induced color enhancement. A commercial ELISA test kit was also used to measure peanut protein in oil spiked with peanut butter. Recovery of peanut residues measured by ELISA was significantly decreased when the peanut butter-spiked oil was heated to 180 degrees C compared to unheated oil. Recovery of spiked peanut butter protein measured by the buffer extraction-colorimetric method was not decreased in heated oil. The method developed here could be used to determine protein levels in crude and refined oil, and to assess the potential for allergen cross-contact from reused cooking oil.

A soybean protein fraction was prepared from defatted soybean seed flour and digested with 29 kinds of commercially available protease originating from preparations of animals, plants, and microorganisms. Some digests, in particular, Ro-digest prepared using a Rhizopus oryzae neutral protease preparation (Peptidase R), displayed strong mitogenic activity toward C3H/HeN mouse spleen cells. The number of spleen CD11b+, CD49b+, interleukin (IL)-12+CD11b+, and interferon (IFN)-gamma+CD49b+ cells significantly increased when cultured with Ro-digest. Similarly, the number of spleen IFN-gamma+CD4+ cells significantly increased in the presence of Ro-digest while that of spleen IL-4+CD4+ cells was largely unchanged. Additionally, 5-week-old male C3H/HeN mice were given diets consisting of ovalbumin (OVA) alone (control diet) or a mixture of OVA and Ro-digest (Ro-digest-added diet) as a protein source for 5 weeks, and the immune properties of the mice were investigated. The number of IL-12+CD11b+ cells was greater in spleens from mice given the Ro-digest-added diet than in those given the control diet. The cytotoxic activity of spleen cells toward the human erythroleukemia cell line, K562, was significantly higher in mice given the Ro-digest-added diet than in those given the control diet. Furthermore, in a microarray analysis of mRNAs extracted from mice Peyer's patch cells, gene expression related to innate immune responses was increased in mice given the Ro-digest-added diet. These results indicate that the Ro-digest might stimulate cellular immune systems, in particular, an innate immunity in mice.

Food enteropathies involve uncontrolled or hypersensitivity reactions to ingested nutrients and may result in IgE and T-helper type 2 (Th2) responses as in food allergy. However, the precise role of B cells in the development of food enteropathies remains uncertain. In this work, we used B cell-deficient mice (B KO) and a model of peanut sensitization to examine the involvement of B lymphocytes in the pathogenesis of food allergy. Results showed that priming of wild-type (WT) mice with peanut proteins induced specific IgG1 and IgE responses in serum, with edema, tissue destruction, epithelial exulceration and inflammatory infiltrate in the gut of sensitized and challenged (S + Peanut) WT animals. In contrast, there was no sera immunoglobulin detection and absence of tissue destruction in the gut of B KO mice, which presented moderate inflammatory infiltrate and villous enlargement after peanut challenge. These animals presented marked decrease in IL-4 and TNF-alpha and high levels of IL-10, TGF-beta, IL-12p40 and IFN-gamma mRNA in the gut. Moreover, the expression of CCL5, CCL11 and CXCL1 was reduced in the gut of B KO mice, in contrast to elevated messages of CCL2 or similar detection of Th1-related chemokines in S + Peanut WT mice. Finally, we provided evidence that B cells are necessary to the development of food-related enteropathies and induction of gut inflammation during allergic reactions to food.

It is suggested that patients with oral allergy syndrome (OAS) respond to pepsin-sensitive allergens, and systemic reactors identify pepsin-resistant allergens. We sought to assess the digestibility of kiwifruit proteins in simulated gastric fluid (SGF), and to compare the immunogenicity of the digests in patients with isolated oral and systemic reactions to kiwifruit. In addition, the effect of pH on digestibility of kiwifruit proteins was investigated. The in vitro resistance of kiwifruit proteins to digestion was determined using SGF. G-immunoglobulin (IgE) binding to digested proteins was investigated by Western blotting using sera from children and adults (aged 5-72 yr) with systemic reactions and patients with isolated oral symptoms. To determine whether pH conditions influence digestion of kiwifruit extracts, digestion at pHs 1.5-7 were compared by SDS-PAGE. Patients with systemic reactions showed IgE binding to digestion-resistant allergens, but patients with oral symptoms reacted only to digestion-labile allergens. An increase in pH from 1.5 to 2.5 significantly reduced pepsin breakdown of kiwifruit allergens. Immunoreactive digested protein fragments were detectable by immunoblot but not Coomassie stain. This study confirms a difference in the lability of food allergens recognized by patients with systemic reactions and those with OAS. Pepsin digestion of kiwifruit proteins was impaired by hypoacidic conditions suggesting that patients with hypoacidic gastric conditions are at increased risk of systemic absorption of allergens. The data indicate that commonly used methods for predicting allergenicity of novel proteins using Coomassie stains may be flawed.

The study reports a case of "pure" tomato allergy in an adult female. The responsible allergen was partially characterized by immunoblot analysis, pepsin digestion, and heating. It had a molecular weight of about 9 kDa and was heat-labile and pepsin-resistant, thus confirming the clinical history. Unfortunately, due to the failure of both 2-dimensional electrophoresis analysis and N-terminal sequencing experiments, it was not possible to characterize the protein further. Based on a comparison with currently known tomato allergens, this seems to be a novel allergen protein.

Plant food allergens belong to a rather limited number of protein families and are also characterized by a number of biochemical and physicochemical properties, many of which are also shared by food allergens of animal origin. These include thermal stability and resistance to proteolysis, which are enhanced by an ability to bind ligands, such as metal ions, lipids, or steroids. Other types of lipid interaction, including membranes or other lipid structures, represent another feature that might promote the allergenic properties of certain food proteins. A structural feature clearly related to stability is intramolecular disulfide bonds alongside posttranslational modifications, such as N-glycosylation. Some plant food allergens, such as the cereal seed storage prolamins, are rheomorphic proteins with polypeptide chains that adopt an ensemble of secondary structures resembling unfolded or partially folded proteins. Other plant food allergens are characterized by the presence of repetitive structures, the ability to form oligomers, and the tendency to aggregate. A summary of our current knowledge regarding the molecular properties of food allergens is presented. Although we cannot as yet predict the allergenicity of a given food protein, understanding of the molecular properties that might predispose them to becoming allergens is an important first step and will undoubtedly contribute to the integrative allergenic risk assessment process being adopted by regulators.

Background: Infantile food protein-induced enterocolitis syndrome (FPIES) is a severe, cell-mediated gastrointestinal food hypersensitivity typically provoked by cow's milk or soy. Solid foods are rarely considered a cause., Objective: To describe the clinical characteristics and natural history of FPIES provoked by solid foods., Methods: Patients with FPIES induced by solid foods were identified and their clinical course compared with a control group with FPIES caused by cow's milk and/or soy evaluated over the same time period., Results: Fourteen infants with FPIES caused by grains (rice, oat, and barley), vegetables (sweet potato, squash, string beans, peas), or poultry (chicken and turkey) were identified. Symptoms were typical of classical FPIES with delayed (median: 2 hours) onset of vomiting, diarrhea, and lethargy/dehydration. Eleven infants (78%) reacted to >1 food protein, including 7 (50%) that reacted to >1 grain. Nine (64%) of all patients with solid food-FPIES also had cow's milk and/or soy-FPIES. Initial presentation was severe in 79% of the patients, prompting sepsis evaluations (57%) and hospitalization (64%) for dehydration or shock. The diagnosis of FPIES was delayed, after a median of 2 reactions (range: 2-5). Thirty patients with typical cow's milk- and/or soy-FPIES were identified for comparison. Overall, 48% of the 44 infants with FPIES were reactive to >1 food protein, and the risk for multiple food hypersensitivity approached 80% in the infants with solid food or soy-induced FPIES. None of the patients developed FPIES to maternally ingested foods while breastfeeding unless the causal food was fed directly to the infant., Conclusions: Cereals, vegetables, and poultry meats, typically regarded as of low allergenic potential, must be considered in the evaluation of FPIES, particularly in infants previously diagnosed with FPIES to cow's milk or soy, and as an initial cause in patients who have been exclusively breastfed. Infants with FPIES are at risk for multiple dietary protein hypersensitivities during an apparent period of immunologic susceptibility. Pediatricians should consider FPIES in the differential diagnosis of shock and sepsis.

Knowledge about cross reactivities among food proteins of plant or animal origin is getting more and more important. Cross reaction provoking often clinical symptoms may be based on close taxonomical relations or on the presence of "archaic", ubiquiter protein structures preserved during phylogenesis. The most significant cross reactivities characterized by increasing incidence are between pollens and foods of plant origin and between latex and fruits.

Allergenic components of cowpea vegetable green seeds (Vigna sinensis) were isolated based on solubility, isoelectric precipitation and molecular mass. The allergenicity of the cowpea fractions was monitored by enzyme-linked immunosorbent assay (ELISA) and skin-prick test. The allergenic albumin fraction was characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and IgE-specific immunoblotting. The 41 and 55 kDa protein components were found to be major allergens and the allergenicity was resistant to heat and proteolytic enzyme digestion. This study confirms the presence of potent allergens in cowpea seeds.

Objective: Even though occupational protein contact dermatitis is not an uncommon finding in occupational dermatology, reports about multiple sensitizations are rare. High-molecular-weight substances such as proteins can pass the epidermis only if it is damaged and cause a sensitization. In a high percentage of case, atopic dermatitis might be the cause of this damage but cannot be regarded as the only cause. An interesting case is presented that was carefully worked up., Methods: Scratch, intracutaneous, and prick or prick-to-prick tests with native occupational allergens were performed as single tests. The patient was patch-tested with the European Standard Series. The determination of allergen-specific IgE was performed by ImmunoCAP. The results were subdivided into six classes. In addition, oral provocations with relevant allergens were performed., Results: The skin tests showed positive type I allergies to beef, lamb, horse, and pork meat, to pork and horse blood as well as to rye and wheat flour, raw potato, and pasta. Weak positive reactions could be found for fowl, duck, goose, and turkey in intracutaneous testing. The ImmunoCAP showed elevated specific IgE values for pork meat, raw potato, and rye and wheat flour. The oral provocation did not show any systemic or skin change., Conclusions: This case report demonstrates how an initial case of contact urticaria turns into protein contact dermatitis. It shows that the diagnosis can be made by means of scratch or prick tests with native occupational allergens. The determination of allergen-specific IgE by ImmunoCAP might be helpful, but a negative result does not exclude protein contact dermatitis.

Calves fed soya proteins may develop severe gastrointestinal disorders. Whether these are predominantly associated with particular Ig subclasses and (or) dietary proteins remains unclear. Therefore, antibody responses to soyabean protein were analysed by dot- and blot-immunobinding in plasma and intestinal mucous secretions. One-month-old calves were fed for 2.5 months liquid diets based on skim milk powder (SMP) or a mixture (2:3, protein basis) of whey and soyabean products including a low antigenic hydrolysed soya protein isolate (HSPI) and a highly antigenic heated soya flour (HSF). Specific antibodies (Abs) of the main isotypes (IgM, IgA, IgG1, IgG2) were characterised by immunostaining of samples which had been previously incubated with nitrocellulose sheets coated with SMP, HSPI or HSF extracts. Plasma collected before feeding experimental diets showed very little specific Abs. By contrast, 2.5 months later, a three-fold increase (P < 0.05) in IgG1 and IgA titres against HSF antigens was observed in calves fed HSF compared with those fed the control or HSPI diet. IgG1 immunoblotting revealed many protein bands from soya in the molecular range of 22-32 and 38-42 kDa. Immunorecognition of specific proteins from SMP and HSPI remained low and similar among animal groups. Specific IgM, IgA and IgG1 titres against HSF, and to a lesser extent HSPI, were significantly higher (P < 0.05) in jejunal mucous secretion of calves fed HSF compared with other groups. Secretions from calves fed HSF bound to many soyabean proteins in the range of 17-23 and 26-38 kDa, with similar patterns for IgA and IgG1. By contrast, only weak bands were found for IgM and IgG2 in all groups of calves. Thus, calves fed antigenic HSF do present specific Abs including IgG1 and IgA isotypes, both systemically and locally. Therefore, IgG1 and (or) IgA rather than IgM and IgG2 Abs may be preferred for assessing the immunogenicity of soyabean products in calves. Interestingly, soyabean immunogenicity was drastically reduced by adequate proteolysis.

Six monoclonal antibodies against Bet v I, the major cross-reactive allergen of birch pollen (Betula verrucosa), were obtained. Four did not react with fruits, but two monoclonal antibodies (mAbs) (5H8 and 9C11) were reactive with apple and other fruits. These two cross-reactive antibodies reacted with identical or overlapping sites, but differed in their relative degree of cross-reactivity toward various fruits and hazelnut. Cross-reactive human IgE antibodies reacted with a nonoverlapping epitope, as indicated by results of a two-site radioimmunoassay (RIA) with the fruit-reactive mAb 9C11. By isoelectric focusing (IEF) in conjunction with immunoblotting, a maximum of seven isoforms could be distinguished. Depletion of birch-pollen extract for Bet v I with the most reactive mAb (7F7) removed approximately 95% of the IgE cross-reactivity between birch pollen and apple extract. The remaining 5% cross-reactive material was still capable of inhibiting the binding of IgE to apple allergen completely, and was reactive with mAbs 5H8 and 3C4. By means of IEF/immunoblot, it was shown that these mAbs recognize an isoform of Bet v I that is poorly, if at all, recognized by mAb 7F7. These results illustrate the heterogeneity of Bet v I, both with respect to the cross-reactive sites as well as to the backbone structure. This type of heterogeneity has possible implications for the use of monoclonal antibodies in allergen standardization.

By a sandwich enzyme-linked immunosorbent assay, a soybean major allergen, Gly m Bd 30K, in soybean products was measured. The allergen occurred at high concentrations in soy milk, tofu, kori-dofu, and yuba, but its content in kinako was small. No allergen was found in fermented foods such as miso, shoyu, and natto. The allergen was clearly shown to occur in meat balls, beef croquettes, and fried chicken that contained soybean protein isolate.

Quantitative analysis and immunohistochemical studies of small intestinal mucosa were performed to investigate the mechanism of mucosal damage in 10 patients with food-sensitive enteropathy. Jejunal biopsy specimens were taken before and after treatment and after clinical relapse following a challenge test. The low villous height of untreated patients normalized after introduction of an elimination diet but declined again to subnormal level after a challenge test. Several other types of cells were significantly increased in the untreated patients in comparison to controls. These included HLA-DR+ (DR+) CD4+ cells in the lamina propria and intraepithelial CD8+ cells. Moreover, those cell patterns, such as increased DR+ CD4+ cells and CD8+ cells, normalized with treatment but regressed to pretreatment levels when the patients were challenged. These findings suggest that activated CD4+ cells in the lamina propria of the small intestinal mucosa, probably by releasing cytokines, may play an important role in contributing to mucosal damage in patients with food-sensitive enteropathy.

Milk replacers containing skim milk powder or a mixture of whey and soy protein provided by a hydrolyzed soy protein isolate or a heated soybean flour were fed to 1-mo-old calves for 101 d. The isolate and the flour provided 56 and 72% of dietary proteins, respectively. Digestibility of feed constituents was measured between 9 to 14 d and 65 to 70 d of treatment. Digestive function was evaluated by measurement of ruminal pH, plasma kinetics of triglyceride and glucose concentrations, and xylose absorption. Antibody production was also recorded. Growth and carcass characteristics were satisfactory for the control diet and the diet based on soy isolate, partly because of high apparent digestibility of protein and lack of antibody synthesis in response to soy isolate. In contrast, protein from soybean flour was poorly digested and highly immunogenic. Ruminal pH at 2.5 h after the meal was unaffected by dietary treatments. Postprandial changes in concentrations of triglycerides and glucose in plasma suggested a lack of abomasal clotting with both diets based on soy. Xylose concentration in plasma was only slightly affected by dietary treatment and calf age. Hydrolyzed soy protein is suitable for veal calves and can account for at least half of protein intake.

The development of local and systemic immune responses to soybean proteins was investigated in early-weaned pigs. Pigs were given either antigenic (ASP, n = 10 pigs) or non-antigenic (NASP, n = 8 pigs) soybean products (6 g of protein/d) from d 5 to 9 of age by stomach tube. After weaning at d 21, pigs were fed diets containing the corresponding soybean products and slaughtered between d 28 to 30. Diarrhea was 2.4-fold more frequent, the size of duodenal villi was reduced by 24 to 36%, and the eosinophil density in the duodenal mucosa was 13 times greater (P < .02) in the ASP pigs compared with the NASP pigs. A larger erythema area (P = .006) was observed in the ASP group than in the NASP group 30 min after an intradermal injection of glycinin, but no significant difference could be detected with alpha- or beta-conglycinin or whole soybean extracts. No difference in skin fold thickness was apparent between groups 24 h later. Intestinal, mesenteric lymph node, and blood lymphocytes did not proliferate when cultured with soybean proteins, regardless of dietary treatment. By ELISA, no difference between groups was observed in the circulating levels of total immunoglobulins (Ig) and IgM. Immunoblotting patterns of raw soybean with sera from 28-d-old pigs showed two bands (22 and 36 kDa) recognized by IgA and IgM, respectively, in the ASP group only. Finally, the adverse effects observed with antigenic soybean flour can be overcome by the use of alcohol-treated products.(ABSTRACT TRUNCATED AT 250 WORDS)

The antibody response to a dietary antigen (soya-bean protein) and the development of oral tolerance was studied in pigs in a family pen system where the piglets are left with their mothers and gradually wean themselves onto a soya-bean-based diet over a 12 week period. In the first experiment three groups of pigs (eight pigs/group) aged either 2, 8 or 13 weeks were immunized with soya-bean protein or ovalbumin (OvA; controls) intra-peritoneally (i.p.) in Quill A adjuvant and subsequently boosted 2 weeks later. All groups showed an IgG response to the injected antigens indicating lack of tolerance induction to the dietary antigen. Interestingly the groups injected with OvA showed an almost identical response to soya-bean protein as the groups injected with soya-bean protein. In a second experiment with a similar protocol, soya bean was withdrawn from the feed before immunization which resulted in lack of response to soya-bean protein in the groups injected with OvA and a lack of response to injected soya-bean protein in the 14-week-old group, indicating that systemic tolerance was established by 12 weeks of age. The results from the two experiments suggest a compartmentalized response to soya-bean protein i.e. local antibody production to dietary soya bean along with systemic tolerance to injected soya-bean protein. The work also suggests that delayed 'natural' weaning may avoid damaging hypersensitivity reactions associated with early weaning.

The soybean protein isolate used in powdered soybean formula is hydrolyzed more extensively than the isolate which is used in liquid soybean formula in most commercial soybean formulas. Previous in vitro studies have shown differences in human antibody response to these soybean protein isolates. Therefore, a prospective clinical study was undertaken to determine if there were differences in adverse reaction rates to these soybean protein isolates. Forty-three patients with possible milk- and/or soy-protein enterocolitis were enrolled in this study. Patients had 3 separate oral food challenges; using milk formula, soybean powder formula and soybean liquid formula. Ten (23%) patients challenged with milk had positive challenges. Fourteen (33%) patients challenged with powdered soy formula had positive challenges while thirteen (30%) challenged with liquid soy formula had positive challenges. In the 10 patients with positive milk challenges, 6 (60%) had a positive soy challenge. In the group with positive soy challenges, 5 reacted to the powdered soy challenge done first, but not the second challenge with the liquid soy formula, and 4 patients reacted to the liquid soy formula challenge done first, but not the second challenge with the powdered soy formula. These results indicate that a significant number of patients with milk protein enterocolitis have soy protein enterocolitis. In addition, an order effect can be demonstrated in the soy challenges because of the tendency to react to the first soy challenge regardless of the type of isolate. These results suggest that a local immune effect caused by the protein may be present.

The weaning of piglets onto soya proteins at 3 weeks old normally results in an active response to the fed protein, as determined by the appearance of serum IgG antisoya antibody. This system thus allows the effects of manipulation on the response to a fed protein to be studied. In animals previously given 1 g of soya protein at birth, the magnitude of the antibody response to soya fed at 3 weeks was decreased, although similar amounts of the fed protein could be detected in serum. In addition, the relative affinity of the dominant interaction between antigen and antibody was reduced in these piglets by almost an order of magnitude. By comparison, the ability of piglets given soya at birth to respond to injected soya was not significantly reduced. These results indicate that the regulation of responses to fed and systemic antigens is largely separate. Very early oral exposure to antigen may affect the ability of neonatal animals to mount immune responses to, specifically, fed proteins while leaving the response to systemic antigen largely intact.

A mouse monoclonal antibody in ascitic fluid or in purified form was subjected to heating or repeated freeze/thaw cycles. Heating the antibody at 80 degrees C for 10 min completely destroyed its ability to bind to antigen. Unpurified antibody in ascitic fluid or purified antibody in buffer was frozen to -196 degrees C in liquid nitrogen and then thawed at room temperature. Purified antibody was also slowly frozen at -4 degrees C followed by thawing at room temperature. Regardless of the manner of freeze/thaw treatment or whether the antibody was purified or not, immunologic competence for its antigen was not lost. The results call into question the belief that freezing and thawing in all cases are necessarily deleterious to antibody structure and function.