Allergen Extracts Research Articles

360° approach to the patient with mite allergy: from scientific evidence to clinical practice.

In the recent years, several important advances have been made in the diagnosis of allergy using molecular techniques. The aetiological diagnosis of allergy using molecular components of allergens allows a more precise definition of the patient's IgE repertoire. Precision medicine is a structural model aimed at personalising healthcare and places the patient at the centre of the specialist's decision-making process. To this end, an accurate characterisation of the external exposome at a molecular level and their putative role as clinically relevant allergens is essential to elucidate the phenotypic diversity of atopic disease, with a view to personalising diagnosis and therapy. It has been proposed a decision algorithm, the Top-Down approach, where the clinical history is set first and is followed by the use of skin tests or specific IgE techniques, which facilitates the clinicians to make decisions. The therapeutic intervention driven by the standard diagnostic approach, but supported by these innovative tools, can lead to a better phenotyping of highly complex patients, and a more appropriate prescription of AIT. To this end, the allergen extracts used for diagnosis require to be of proven quality and contain the most relevant allergens. Likewise, allergen vaccines must gather efficacy, safety, duration, and patient compliance, hence the demand for new vaccines to overcome these drawbacks.

IgE Antibody Assay-Based Allergen Database(CLSI-IL/A-37)

There are 8 established allergen databases (IUIS/WHO ANDB, Allergome, AllFam, AllergenOnline, SDAP, COMPARE, IEDB and AllerBase).None of these provide a comprehensive listing of the internationally-accepted nomenclature codes and Linnean system descriptors of the~1000 extract and molecular allergens used world-wide in diagnostic single-and multi-plex IgE antibody assays. The new Diagnostic Allergen DataBase (DADB) has been prepared by an international scientific committee of the Clinical and Laboratory Standards Institute to specifically provide these data together in a readily-searchable Excel database. Using peanut as an example, the DADB lists the allergens’ unique assay working codes (F13), general category (food), principal IgE antibody assay method(s) where the allergen specificity is available, common name (peanut), taxonomical name (Arachis hypogaea), and the NCBI, NPU, LOINC, Allergome and IUIS/WHO descriptor codes for allergens and their assays. The DADB benefits manufacturers of allergen-specific IgE assays by providing unambiguous specificity descriptors. With the DADB, allergen extract manufacturers will have clearly defined specificity targets for allergenic product development. Government regulators will have clarity on the specificity of new allergen reagents that are being submitted for clearance. IgE antibody proficiency testing programs (e.g. CAP) will be able to clearly define the IgE antibody specificities being measured in their challenge sera. The clinician will precisely know the allergen specificity of IgE antibody that is measured in their reference laboratory. Finally, the allergic patient will be assured that their allergic sensitization is being assessed in an assay with a well-characterized allergen reagent.

Advancements in Allergen Immunotherapy for the Treatment of Atopic Dermatitis.

Atopic dermatitis (AD) is a chronic inflammatory skin condition that affects individuals of all age groups, manifesting as a spectrum of symptoms varying from mild to severe. Allergen immunotherapy (AIT) involves the administration of allergen extracts and has emerged as a potential treatment strategy for modifying immune responses. Its pathogenesis involves epidermal barrier dysfunction, microbiome imbalance, immune dysregulation, and environmental factors. Existing treatment strategies encompass topical steroids to systemic agents, while AIT is under investigation as a potential immune-modifying alternative. Several studies have shown reductions in the severity scoring of atopic dermatitis (SCORAD) scores, daily rescue medication use, and visual analog scale (VAS) scores following AIT. Biomarker changes include increased IgG4 levels and decreased eosinophil counts. This review provides valuable insights for future research and clinical practice, exploring AIT as a viable option for the management of AD.

Characterization of Pollen Grain and Pollen Extract from Common Allergenic Plants in the Philippines

Allergic rhinitis and asthma are prevalent in tropical countries like the Philippines. Pollen grains from anemophilous plant species may trigger these respiratory allergies. Pollen extracts from common allergenic plants are currently produced locally in the Philippines and are utilized for the diagnosis and immunotherapy of patients with respiratory allergies. Further improvement of their quality is necessary to avoid false negative or false positive diagnoses, as well as to increase the effectiveness of immunotherapy and avoid adverse reactions. This study aimed to characterize the morphology of the collected pollen grains and assess the pollen purity, protein concentration, and electrophoretic profile of the allergenic pollen extracts. Pollen identification and percent purity of the pollen collections were assessed via microscopy and hemocytometer. Lowry assay was performed to identify the total protein concentration of the pollen extracts before subjecting them to SDS-PAGE and silver staining. The pollen samples from varying plant families have distinct features that will aid in their proper identification. Collected aggregates of pollen samples with pollen purity of < 90%, like T. procumbens, A. saman, L. leucocephala, and M. indica, manifest darker colors compared to their actual color. On the other hand, the color of collected pollen samples with pollen purity ≥ 90% are commonly yellow or off-white, which is their natural color. The total protein concentration of the crude pollen extracts ranges from 2.87–9.63 mg/mL. The pollen extracts revealed protein bands at molecular weights ranging from 10–250 kD on SDS-PAGE. Proper characterization of a particular pollen may help in assessing the purity of the collected pollen. Higher pollen purity may contribute to the assurance that the positive diagnosis was due to the target pollen of a plant and not due to contaminations from other components. The result of this study may be considered for the improvement of the crude pollen allergen extract production for clinical use. It is also recommended to assess the effect of the environmental condition where the pollen was collected in terms of the actual proteins present after allergen extraction.

Diagnostic gap due to missing patch test allergens—status quo and possible scenarios for mitigation

SummaryAccording to European Directive 2001/83/EC, test and therapeutic allergens are medicinal products in all Member States of the European Union. This applies equally to prick test and to patch test (PT) allergens (haptens). All test allergens commercially marketed in Germany are finished medicinal products requiring marketing authorization (MA). Currently, 211 PT substances are authorized in Germany, and an additional 59 are in an ongoing MA process and are marketable under a transitional provision until a decision on MA is made. The regulatory guidance (CMDh/399/2019) of the Co-ordination Group for Mutual Recognition and Decentralized Procedures—Human (CMDh), published in July 2020, specifies the regulatory requirements for different allergen products. Due to differences in origin and exposure, use, mode of action, and safety risks, the guideline clearly differentiates between products with active ingredients of biological origin (allergen extracts from natural source materials) and products with active ingredients of non-biological origin (hapten-based PT substances). Currently, guideline-compliant patch testing is hampered by the lack of numerous commercial PT allergens from the standard and special test series. Background and possible scenarios for mitigation are presented here.

Correlation of Blomia tropicalis-specific immunoglobulin epsilon profiles with family history of atopy in a Filipino population.

House dust mites are the major source of indoor allergens in the tropical and subtropical regions with Blomia tropicalis (Bt) allergens as one of the leading causative agents of sensitization among patients from the tropics. Despite the clinical importance of Bt in various populations, its allergenicity remains unclear among Filipino allergic patients. This study determined the sensitization profiles of allergic Filipinos against Bt allergens and its correlation with atopy. Total immunoglobulin epsilon (IgE) (n = 960), Bt-specific IgE (n = 247), and Blomia tropicalis 5 (Blo t 5)-specific IgE (n = 87) profiles of allergic and nonallergic subjects were measured through enzyme-linked immunosorbent assay (ELISA). Point-biserial correlation coefficient was used to determine the association between Bt-specific IgE levels and selected demographics. Inhibition ELISA was performed to measure the inhibition capacity of recombinant Blo t 5 (rBlo t 5) against Bt allergen extracts. Mean total IgE levels of allergic cases (n = 171) were significantly higher (P < 0.001) compared to the mean IgE levels of nonallergic controls (n = 76). Among allergic subjects, 58% were sensitized to Blo t extract and 80% of which were sensitized to rBlo t 5 allergen. A positive correlation was observed between Bt-specific IgE and family history of atopic disease (P = 0.031). Inhibition assay revealed that 54% mean reactivity of 7 plasma samples was caused by rBlo t 5, validating that rBlo t 5 is a major allergen in Bt. This study has shown the importance of Bt as an allergen source that sensitizes atopic Filipino subjects. Hence, inclusion of Bt allergen extract and rBlo t 5 in the panel for allergy diagnosis and immunotherapy in Filipino populations is strongly recommended.

The Sensitization Profile for Selected Food Allergens in Polish Children Assessed with the Use of a Precision Allergy Molecular Diagnostic Technique.

Individual populations show a variety of sensitization patterns, which may be associated with the geographic region, climate, dietary habits, or ways of preparing food. The purpose of this study was to comprehensively assess the food allergy sensitization profile in Polish children, particularly to eight food allergens (so-called "the Big 8"): cow milk, eggs, wheat, soybeans, fish, crustacean shellfish, tree nuts, and peanuts. To assess the prevalence and serum levels of specific immunoglobulins E (sIgE), we analyzed the results obtained from selected laboratories located in all regions of Poland that used the multiplex ALEX® test in the period from 2019 to 2022. Results from 3715 children were obtained. The mean age of the study population was 7.0 years. The results were stratified by age: <12 months (3.63%), 1-5 years (39.54%), 6-13 years (46.32%), and 14-18 years (10.0%). The final analysis included the sIgE results obtained with 95 food extracts and 77 food allergen molecules. The highest rates of sIgE to food allergen extracts were found for peanut (29.20%), hazel (28.20%), and apple (23.60%), and those to allergenic molecules were found for the PR-10 family of molecules (Cor a 1.0401 (23.77%), Mal d 1 (22.37%), Ara h 8 (16.93%), and globulin 7/8S (Ara h 1; 15.59%)). The lowest rates of sIgE reactivity to extracts were found for strawberry (0.40%), oregano (0.30%), and thornback ray (0.16%), and those to allergenic molecules were found for Mal d 2 (0.27%) (thaumatin-like protein, TLP), Ani s 1 (0.30%) (Kunitz-type serine protease inhibitor), and Che a 1 (0.43%) (Ole e 1 family). The rates of sensitization to storage proteins of the analyzed "the Big 8" molecules decreased significantly (p < 0.05) with age. Conversely, the rates of sensitization to PR-10 family proteins increased significantly with age. The three most common allergens in Poland, regardless of whether IgE was assayed against extracts or molecules of food allergens, were peanut, hazel, and apple (in different order depending on the ranking). A detailed analysis of sensitization to the extracts and molecules of main food allergens based on the results of a multiplex ALEX® test demonstrated the sensitization profile in Polish children (including molecular sensitization, particularly the "the Big 8" food allergen molecules), which shows considerable differences in comparison with those in other countries. Serum sIgE analysis of children from all regions of Poland revealed a food allergen molecular sensitization profile that changes with age.

Novel monoclonal antibodies against house dust mite allergen Der p 21 and their application to analyze allergen extracts.

Allergen extracts and recombinant allergens are used in allergy diagnostics and immunotherapy. Since allergen extracts from different manufacturers lack proper standardization regarding their composition, monoclonal antibodies (MAbs) against specific allergen components can be used for their identification and quantification in allergen extracts. This study aimed to generate MAbs against allergen Der p 21 of Dermatophagoides pteronyssinus for the analysis of allergen extracts. Recombinant Der p 21 was expressed in E. coli and purified using affinity chromatography. MAbs against Der p 21 were generated using hybridoma technology. House dust mite (HDM) allergen extracts were analyzed using the newly developed sandwich enzyme-linked immunosorbent assay, Western blotting and microarray immunoassay. MAbs raised against recombinant Der p 21 were characterized in detail and proven to be reactive with natural Der p 21. Highly specific sandwich enzyme-linked immunosorbent assay for the quantification of Der p 21 was developed and optimized. The allergen was detected and its concentration was determined in only three of six analyzed HDM allergen extracts from different manufacturers. HDM analysis by MAb-based immunoassays shows their differences in allergen composition. The results demonstrate the importance of allergen-specific MAbs as a tool for the characterization of allergen extracts and the need for their appropriate standardization before their use for allergy diagnostics or immunotherapy.

To stay or not to stay intact as an allergen: the endolysosomal degradation assay used as tool to analyze protein immunogenicity and T cell epitopes.

Antigen uptake and processing of exogenous proteins is critical for adaptive immunity, particularly for T helper cell activation. Proteins undergo distinct proteolytic processing in endolysosomal compartments of antigen-presenting cells. The resulting peptides are presented on MHC class II molecules and specifically recognized by T cells. The in vitro endolysosomal degradation assay mimics antigen processing by incubating a protein of interest with a protease cocktail derived from the endolysosomal compartments of antigen presenting cells. The kinetics of protein degradation is monitored by gel electrophoresis and allows calculation of a protein's half-life and thus endolysosomal stability. Processed peptides are analyzed by mass spectrometry and abundant peptide clusters are shown to harbor T cell epitopes. The endolysosomal degradation assay has been widely used to study allergens, which are IgE-binding proteins involved in type I hypersensitivity. In this review article, we provide the first comprehensive overview of the endolysosomal degradation of 29 isoallergens and variants originating from the PR-10, Ole e 1-like, pectate lyase, defensin polyproline-linked, non-specific lipid transfer, mite group 1, 2, and 5, and tropomyosin protein families. The assay method is described in detail and suggestions for improved standardization and reproducibility are provided. The current hypothesis implies that proteins with high endolysosomal stability can induce an efficient immune response, whereas highly unstable proteins are degraded early during antigen processing and therefore not efficient for MHC II peptide presentation. To validate this concept, systematic analyses of high and low allergenic representatives of protein families should be investigated. In addition to purified molecules, allergen extracts should be degraded to analyze potential matrix effects and gastrointestinal proteolysis of food allergens. In conclusion, individual protein susceptibility and peptides obtained from the endolysosomal degradation assay are powerful tools for understanding protein immunogenicity and T cell reactivity. Systematic studies and linkage with in vivo sensitization data will allow the establishment of (machine-learning) tools to aid prediction of immunogenicity and allergenicity. The orthogonal method could in the future be used for risk assessment of novel foods and in the generation of protein-based immunotherapeutics.

Сенсибилизация к эпидермальным аллергенам у детей с аллергопатологией: 30-летний мониторинг

Aim: to analyze changes in sensitization to common epidermal allergens over time in children with allergic diseases in a metropolitan area. Patients and Methods: summary and analysis of 26,343 skin prick test protocols of children aged 4–18 years who were examined and treated between 1989 and 2019 were performed. A standard skin scarification test was used to determine an individual's sensitivity to common epidermal allergens, including feline, canine, sheep, and equine hair/epidermis, pillow feathers, and human hair, using a drop of water-salt allergen extract. To calculate the average exponents, i.e., the annual level and severity of sensitization, visual scoring (+1, +2, +3, +4) was used. This was done by calculating the percentage ratio of the number of patients with hypersensitivity to the allergen to the total number of patients examined for this allergen during a particular year. The proportion of low (1-2) and high (3-4) scores were also compared. Results: the cohort of allergic children was followed up for 30 years, during which there was a constant increase in IgE-hypersensitivity to epidermal allergens. Specifically, there was an 8.5-fold increase in sensitivity to feline hair/epidermis, with a 27.5-fold increase in the proportion of children scoring 3–4 points. Sensitivity to canine hair/epidermis increased by 5.2 times, while sensitivity to horse dander increased by 24%, sheep's hair by 23.7%, pillow feather by 2.7 times, and human hair by 2.8 times. Meanwhile, a retrospective analysis has shown a slowdown in the rate of increase in the number of sensitized patients and a decrease in the severity of specific hypersensitivity, mainly in the range of 1–2 points, over the last 15 years. Conclusion: the retrospective analysis indicates a progressive increase in epidermal sensitization to all studied allergens, with a general increase in the severity of hypersensitivity. This suggests an increase in close direct and indirect contacts with animals, as well as the potential effect of aggressive urban ecology on the immune balance of a growing child and the transformation of immunogenicity of animal allergens themselves. Given this, it is important to consider the efficacy of elimination measures and the choice of effective methods for preventing epidermal allergy. This includes the formation of immune tolerance to ensure reliable and long-term protection. KEYWORDS: animal allergens, epidermal allergy, children, sensitization monitoring, increased hypersensitivity; perspectives. FOR CITATION: Bardenikova S.I., Lokshina E.E., Dovgun O.B., Shavlokhova L.A., Bogdanova N.A., Serebrovskaya N.B., Mstislavskaya S.A., Kuznetsov G.B. Sensitization to epidermal allergens in children with allergic disorders: a 30-year follow-up study. Russian Medical Inquiry. 2024;8(3):118–123 (in Russ.). DOI: 10.32364/2587-6821-2024-8-3-1.

Procedure for a standardized preparation of skin prick test solutions for the diagnosis of occupational type I allergies in the absence of commercial extracts.

In order to ensure valid diagnostics for occupational test allergen solutions despite the ongoing reduction in the availability of commercial test extracts, a plan B was initiated for the possible production of skin prick test (SPT) solutions in public pharmacies. For important occupational allergen sources (wheat and rye, storage mites, animal epithelia, mold material) laboratory extraction methods were analyzed in comparison to pharmacy compatible extraction methods regarding protein quantity and quality in SDS-PAGE combined with silver staining. Subsequently, using the example of bovine epithelia, adapted extraction procedures as well as in-process and final product controls were transferred to a public pharmacy. Allergen sources with a high protein content, such as wheat and rye grains as well as storage mites, showed good comparability of the extractable protein quantity and protein pattern, regardless of the applied extraction method. In contrast, allergen source materials with a low total protein content, such as animal epithelia and molds, can benefit from laboratory extraction conditions such as mechanical disruption and specific buffer additives. In the qualitative protein silver staining, characteristic protein patterns were identified for each allergen source. Depending on the extraction method, only minor differences in total protein patterns were observed in animal epithelia and molds. Using source materials from two suppliers, the resulting allergen extracts displayed clear differences in protein content in storage mites and quantitative and qualitative differences in molds. A practical preparation attempt of SPT solutions in a public pharmacy was successful. SPT solutions prepared with adapted pharmacy extraction methods showed a comparable protein and Bos d 2 allergen content and equivalent qualities in the protein pattern compared to a previously available commercial SPT solution. Accordingly, it can be assumed that standardized SPT solutions with sufficient allergen quality for occupational allergen sources can be prepared in public pharmacies if certified allergen sources with appropriate protein content are available.

Skin test (Skin Prick Test) in food allergy

Skin tests, also known as prick tests, in food allergies are restricted to IgE-mediated food allergies, with results that barely indicate sensitization to a certain food and do not necessarily suggest food allergy. The clinical context of the patient is the decisive point before performing any type of food allergy skin test; Contextualizing and relating the patient's clinical picture with the immunological mechanism of IgE-mediated allergies will allow a better selection of the allergy test for each case. Positive tests should be interpreted more carefully, and consider that during early childhood it is common that several patients may have negative specific serum IgE tests in the blood for food allergens compared to those performed on the skin, which are positive. Skin testing can be carried out with standardized strata of foods, but they are not always available for all foods and some are very unstable. Skin tests can even be performed with raw foods, using the prick to prick technique to perform the puncture with fresh foods (especially fruits or vegetables) or other products that are not commercially available. The skin prick test is a test where, after having placed the allergenic extract in drops or with fresh food itself, the puncture must be carried out with a standardized lancet.

RAST, Inmunoblot, Immunocap and ISAC in food allergy

skin tests; Sensitization; IgE-mediated allergy; allergenic extract. After the discovery of IgE, technological advances have provided new laboratory tools for the quantification of allergen-specific IgE antibodies in serum and on the surface of basophils-mast cells. In vitro testing offers numerous advantages: accurate quantitation, lack of drug interference, safety, and long-term storage of samples. Quantitative immunoassays for IgE antibodies can be an adjunct to skin testing. The allergen reagent in solid phase (allergosorbent) or liquid is the main component of the assay that confers specificity to the IgE antibody test. It is the most complex and highly variable reagent in IgE antibody assays. The choice to use diagnostic recombinants on a single rather than multiple platforms is made on a case-by-case basis (considering prior history and clinical profile) and in an allergen-dependent manner. Although most food allergies are limited to a small number of possible triggers, these foods are very complex when evaluating their allergenic potential. The possibility of fractionating the allergen and understanding some of its components as potentially important to define the risk of clinical reaction, cross-reactivity, or persistence of allergy, opened a new era in the field of allergy, called molecular allergy. The identification of the allergenic component responsible for the reactions is an important tool to confirm the information and severity of the symptoms, natural history of the disease, possibility of cross-reactivity and clinical symptoms (allergy markers).

T cells specific to multiple Bet v 1 peptides are highly cross-reactive toward the corresponding peptides from the homologous group of tree pollens.

Allergens from Fagales trees frequently cause spring allergy in Europe, North America, and some parts of Asia. The definition of the birch homologous group, which includes birch (Bet v), oak (Que a), alder (Aln g), hazel (Cor a), hornbeam (Car b), beech (Fag s), and chestnut (Cas s), is based on high allergen sequence identity and extensive IgE cross-reactivity. Clinical effect was seen during the alder/hazel, birch, and oak pollen seasons after treatment with tree SLIT-tablets containing only birch allergen extract. Here, we characterize T-cell reactivity with respect to epitope specificities and cross-reactivity toward various Bet v 1 family members, (PR-10/group 1 major allergens). This cross-reactivity may be part of the immunological basis of clinical effect or cross-protection when exposed to birch homologous tree species. T-cell lines were generated from 29 birch-allergic individuals through stimulation of peripheral blood mononuclear cells (PBMCs) with birch/Bet v or oak/Que a allergen extracts. T-cell responses to allergen extracts, purified group 1 allergens, and overlapping 20-mer peptides (Bet v 1, Aln g 1, Cor a 1, and Que a 1) were investigated by T-cell proliferation and cytokine production. Cross-reactivity was evaluated based on Pearson's correlations of response strength and further investigated by flow cytometry using tetramer staining for homologous peptide pairs. T-cell reactivity toward extracts and group 1 allergens from across the birch homologous group was observed for birch/Bet v as well as oak/Que a T-cell lines. T-cell lines responded to multiple Bet v 1 homologous peptides from Aln g 1 and Cor a 1 and a subset of Que a 1 peptides. Significant Pearson's correlations between frequently recognized peptides derived from Bet v 1 and the corresponding peptides derived from alder, hazel, and oak strongly supported the T-cell cross-reactivity toward these allergens. Cross-reactivity between birch and birch homologous peptides was confirmed by pMHCII tetramer staining. T cells from birch tree pollen allergic individuals respond to multiple trees within the birch homologous group in accordance with the level of sequence homology between Bet v 1 family members, (PR-10 allergens) from these allergen sources, confirming the basis for clinical cross-protection.

Cholinergic sensing of allergen exposure by airway epithelium promotes type 2 immunity in the lungs

Nonneuronal cells, including epithelial cells, can produce acetylcholine (ACh). Muscarinic ACh receptor antagonists are used clinically to treat asthma and other medical conditions; however, knowledge regarding the roles of ACh in type 2 immunity is limited. Our aim was to investigate the roles of epithelial ACh in allergic immune responses. Human bronchial epithelial (HBE) cells were cultured with allergen extracts, and their ACh production and IL-33 secretion were studied invitro. To investigate immune responses invivo, naive BALB/c mice were treated intranasally with different muscarinic ACh receptor antagonists and then exposed intranasally to allergens. At steady state, HBE cells expressed cellular components necessary for ACh production, including choline acetyltransferase and organic cation transporters. Exposure to allergens caused HBE cells to rapidly release ACh into the extracellular medium. Pharmacologic or small-interfering RNA-based blocking of ACh production or autocrine action through the M3 muscarinic ACh receptors in HBE cells suppressed allergen-induced ATP release, calcium mobilization, and extracellular secretion of IL-33. When naive mice were exposed to allergens, ACh was quickly released into the airway lumen. Aseries of clinical M3 muscarinic ACh receptor antagonists inhibited allergen-induced IL-33 secretion and innate type 2 immune response in the mouse airways. In a preclinical murine model of asthma, an ACh receptor antagonist suppressed allergen-induced airway inflammation and airway hyperreactivity. ACh is released quickly by airway epithelial cells on allergen exposure, and it plays an important role in type 2 immunity. The epithelial ACh system can be considered a therapeutic target in allergic airway diseases.

T-cell subset changes during the first year of pre-seasonal allergoid allergen-specific immunotherapy

Allergen-specific immunotherapy (AIT) is the only treatment for type I allergy, which achieves long-lasting effects. Repeated subcutaneous applications of allergen extracts cause a protective antibody response and an immune deviation of T cells. In AIT with allergoids, chemically modified allergen extracts are injected. During a so-called special pre-seasonal application scheme, after the initial phase of applying increased doses of allergoids is followed by natural allergen exposure as a maintenance phase. The effectiveness of allergoid vaccines has been described regarding the improvement of clinical symptoms and the development of protective humoral responses.In this longitudinal observational study, we sought to investigate changes at the T cell level in pre-seasonal AIT with allergoid. Different subsets within CD4+ and CD8+ T cells were monitored by flow cytometry in PBMC of patients known to possess protective antibody responses.Compared to before treatment, a small early boost among allergenic Th cells was observed after 4 months of AIT. In line, a slight Th2 bias was observed after 4 months within circulating T follicular T cells, Tfh and Tfc, representing pre-existing memory Th2 cells. Furthermore, it was demonstrated that responsiveness of CD8+ T cells to allergen stimulation decreased during the course of treatment. Apart from that, we found an influence of the meteorological season on the activation profile of Tfh and Tfc over the course of the treatment.Together, this is the first study investigating changes of different T cell subsets over the course of an allergoid AIT against airborne allergens. Our findings match previous reports on conventional AIT, especially the initial increase of Th2 responses. However, the observed changes were less pronounced which may be either due to the modification of allergens or to the reduced maintenance dose provided by natural allergen exposure compared to a perennial protocol.

Allergy immunotherapy for allergic fungal respiratory diseases.

Background: Allergy immunotherapy (AIT) with fungal extracts is not as straight forward as that with other inhalants. The complexities relate to the number of airborne fungal spores, the limited data on the exposure to the spores of individual species of fungi and their clinical importance, the poor quality of the fungal allergen extracts that are available for the diagnosis and treatment, and the lack of controlled studies establishing dosing and efficacy of AIT with fungal extracts except for Alternaria. Objective: The objective was to review what is known with regard to the role of fungi in causing allergic respiratory diseases as well as the evidence that exists for the role of AIT as a treatment for these conditions. Methods: A search was conducted of PubMed, textbooks, known articles on immunotherapy with fungal extracts, and references derived from these primary sources. Results: Nine immunotherapy studies that used Alternaria or its major allergen Alt a 1 and two studies that used Cladosporium herbarum were identified. When a good quality extract was administered in adequate doses, immunotherapy with Alternaria was as effective as that with other inhalant allergens. There was a suggestion of efficacy with a specially prepared Cladosporium extract, but systemic reactions were common and limited the tolerated dose. The use of immunotherapy as an adjunct treatment for allergic fungal sinusitis is briefly reviewed, but controlled trials are lacking. Conclusion: Fungal immunotherapy should largely be limited to Alternaria alternata and perhaps C. herbarum. Under conditions of demonstrated exposure to a particular species of fungus and with symptoms that correlate with that exposure as well as availability of an apparently potent extract of that fungus to which the patient is sensitive that fungus may be considered for immunotherapy. Fungal (mold) mixes should not be used for diagnosis or therapy.

Recent advances in epicutaneous immunotherapy and potential applications in food allergy.

Given the potent immunological properties of the skin, epicutaneous immunotherapy (EPIT) emerges as a promising treatment approach for inducing immune tolerance, particularly for food allergies. Targeting the highly immunocompetent, non-vascularized epidermis allows for the application of microgram amounts of allergen while significantly reducing the risk of allergen passage into the bloodstream, thus limiting systemic allergen exposure and distribution. This makes EPIT highly suitable for the treatment of potentially life-threatening allergies such as food allergies. Multiple approaches to EPIT are currently under investigation for the treatment of food allergy, and these include the use of allergen-coated microneedles, application of allergen on the skin pretreated by tape stripping, abrasion or laser-mediated microperforation, or the application of allergen on the intact skin using an occlusive epicutaneous system. To date, the most clinically advanced approach to EPIT is the Viaskin technology platform. Viaskin is an occlusive epicutaneous system (patch) containing dried native allergen extracts, without adjuvants, which relies on frequent application for the progressive passage of small amounts of allergen to the epidermis through occlusion of the intact skin. Numerous preclinical studies of Viaskin have demonstrated that this particular approach to EPIT can induce potent and long-lasting T-regulatory cells with broad homing capabilities, which can exert their suppressive effects in multiple organs and ameliorate immune responses from different routes of allergen exposure. Clinical trials of the Viaskin patch have studied the efficacy and safety for the treatment of life-threatening allergies in younger patients, at an age when allergic diseases start to occur. Moreover, this treatment approach is designed to provide a non-invasive therapy with no restrictions on daily activities. Taken together, the preclinical and clinical data on the use of EPIT support the continued investigation of this therapeutic approach to provide improved treatment options for patients with allergic disorders in the near future.

The optimisation and standardisation of Indonesian house dust mites allergenic extract as a desensitising agent

Background: The different sequences of the allergen protease can induce different immunogenicity for the development of immunotherapy. Dermatophagoides pteronyssinus that has Der p1 as major protein will be developed as local allergen to be immunotherapy for treating rhinitis allergy. Objective: This study aims to optimize and standardize the local Indonesian House Dust Mites Allergenic Extract from the harvesting into the extraction process. Method: The harvesting media used is corn flour placed in certain conditions to produce the mites. After harvesting the flour in different weeks, the flour was extracted and the lipid content and Der p1 assay were measured. Result: The result of the study is that the 3 weeks-harvesting time came higher for those parameters than the other harvesting time (p &lt; 0.05). Conclusion: Thus, it was concluded that the standardized harvesting and extraction process had been developed to produce the standardised Indonesian House Dust Mites Allergenic Extract.

Sensitivity, specificity and positive predictive value of ALEX2 multiplex examination in patients suffering from atopic dermatitis and reaction to tomatoes

ABSTRACT The aim of the study is to evaluate the clinical reactions to tomatoes in atopic dermatitis (AD) patients, and to analyse the results of specific IgE to allergens of tomatoes and to molecular components of pollen, seeds, fruits, vegetables and latex with the use of ALEX2 Allergy Xplorer test. According to our results, 15% of AD patients suffer from clinical reactions to tomatoes; we confirmed the low sensitivity and low positive predictive value of specific IgE to molecular component Sola l 6 and allergen extract Sola l, the specificity of this test is higher. The significantly higher sensitisation to other nonspecific lipid transfer proteins is observed in patients sensitised to tomato allergens. An elimination diet and an exposure test are recommended to detect a reaction to tomatoes. ALEX2 Allergy Xplorer test gives us a comprehensive picture of sensitisation and the possibility of evaluation of cross-reacting allergens.