AllergoOncology: Biomarkers and refined classification for research in the allergy and glioma nexus-A joint EAACI-EANO position paper.

Epidemiological studies have explored the relationship between allergic diseases and cancer risk or prognosis in AllergoOncology. Some studies suggest an inverse association, but uncertainties remain, including in IgE-mediated diseases and glioma. Allergic disease stems from a Th2-biased immune response to allergens in predisposed atopic individuals. Allergic disorders vary in phenotype, genotype and endotype, affecting their pathophysiology. Beyond clinical manifestation and commonly used clinical markers, there is ongoing research to identify novel biomarkers for allergy diagnosis, monitoring, severity assessment and treatment. Gliomas, the most common and diverse brain tumours, have in parallel undergone changes in classification over time, with specific molecular biomarkers defining glioma subtypes. Gliomas exhibit a complex tumour-immune interphase and distinct immune microenvironment features. Immunotherapy and targeted therapy hold promise for primary brain tumour treatment, but require more specific and effective approaches. Animal studies indicate allergic airway inflammation may delay glioma progression. This collaborative European Academy of Allergy and Clinical Immunology (EAACI) and European Association of Neuro-Oncology (EANO) Position Paper summarizes recent advances and emerging biomarkers for refined allergy and adult-type diffuse glioma classification to inform future epidemiological and clinical studies. Future research is needed to enhance our understanding of immune-glioma interactions to ultimately improve patient prognosis and survival.

Study protocol to investigate the efficacy of confocal laser endomicroscopy-based selective single-elimination diet over standard fivefold elimination diet in patients with endomicroscopically proven food intolerance: app-assisted, monocentric, double-blind, randomised and controlled trial in Germany.

INTRODUCTION: Imprecise nutritional recommendations due to a lack of diagnostic test accuracy are a frequent problem for individuals with adverse reactions to foods but no precise diagnosis. Consequently, patients follow very broad and strict elimination diets to avoid uncontrolled symptoms such as diarrhoea and abdominal pain. Dietary limitations and the uncertainty of developing gastrointestinal symptoms after the inadvertent ingestion of food have been demonstrated to reduce the quality of life (QoL) of affected individuals and subsequently might increase the risk of malnutrition and intestinal dysbiosis. This trial aims to investigate the effects of a tailored diet based on the confocal laser endoscopy (CLE) examination result to limit the side effects of unspecific and broad elimination diets and to increase the patient's QoL. METHODS AND ANALYSIS: The study is designed as a prospective, double-blind, monocentric, randomised and controlled trial conducted at the University Hospital of Schleswig-Holstein, Campus Lübeck, Germany. One hundred seventy-two patients with non-IgE-related food allergies and positive CLE results will be randomised to either a tailored diet or a standard fivefold elimination diet. The primary endpoints are the difference between the end and the start of the intervention in health-related QoL and the sum score of the severity of symptoms after 12 weeks. Key secondary endpoints are changes in the severity of symptoms, further QoL measurements, self-assessed state of health and number of days with a pathologically altered stool. Microbiome diversity and metabolome of stool, urine and blood will also be investigated. Safety endpoints are body composition, body mass index and adverse events. ETHICS AND DISSEMINATION: The study protocol was accepted by the ethical committee of the University of Lübeck (AZ: 22-111) on 4 May2022. Results of the study will be published in peer-reviewed journals and presented at scientific meetings. TRIAL REGISTRATION NUMBER: German Clinical Trials Register (DRKS00029323).

EAACI Molecular Allergology User's Guide 2.0.

Since the discovery of immunoglobulin E (IgE) as a mediator of allergic diseases in 1967, our knowledge about the immunological mechanisms of IgE-mediated allergies has remarkably increased. In addition to understanding the immune response and clinical symptoms, allergy diagnosis and management depend strongly on the precise identification of the elicitors of the IgE-mediated allergic reaction. In the past four decades, innovations in bioscience and technology have facilitated the identification and production of well-defined, highly pure molecules for component-resolved diagnosis (CRD), allowing a personalized diagnosis and management of the allergic disease for individual patients. The first edition of the "EAACI Molecular Allergology User's Guide" (MAUG) in 2016 rapidly became a key reference for clinicians, scientists, and interested readers with a background in allergology, immunology, biology, and medicine. Nevertheless, the field of molecular allergology is moving fast, and after 6 years, a new EAACI Taskforce was established to provide an updated document. The Molecular Allergology User's Guide 2.0 summarizes state-of-the-art information on allergen molecules, their clinical relevance, and their application in diagnostic algorithms for clinical practice. It is designed for both, clinicians and scientists, guiding health care professionals through the overwhelming list of different allergen molecules available for testing. Further, it provides diagnostic algorithms on the clinical relevance of allergenic molecules and gives an overview of their biology, the basic mechanisms of test formats, and the application of tests to measure allergen exposure.

Vegan diets from an allergy point of view - Position paper of the DGAKI working group on food allergy.

Vegan diets are currently attracting a great deal of attention. However, avoiding animal-based foods restricts the diet and is associated with risks, the extent and medical implications of which are at present not sufficiently understood. Elimination diets represent the usual therapeutic long-term management in the presence of food allergy. In order to understand the risks of vegan diets and to discuss these critically from the perspective of food allergies, the expertise of a nutritionist/dietitian with expertise in this area is indispensable. This position paper deals with the incentives behind and the benefits of a plant-based diet. The knowledge required to cover macro- and micronutrient dietary requirements is presented. Using the avoidance of cow's milk as an example, the challenges of adequately meeting nutritional needs are identified and (so-called) milk alternatives are evaluated from an allergy and nutritional point of view. Finally, other plant-based (substitute) products are evaluated from the same perspective, as significant protein sources in vegan diets (e.g., legumes, nuts, and seeds) are at the same time potential and potent triggers of allergic reactions. However, the allergic potential of many substitute products cannot be fully assessed at present due to gaps in research. Wheat as the most important trigger for anaphylaxis in adults is also evaluated. Finally, the increase in ultra-processed products in the (vegan) food sector and their potential consequences for the immune system are discussed.

Food-Induced Anaphylaxis: Data From the European Anaphylaxis Registry.

BACKGROUND: Food is one of the most common elicitors of anaphylaxis, with an increasing incidence over recent years. OBJECTIVES: To characterize elicitor-specific phenotypes and identify factors enhancing the risk or severity of food-induced anaphylaxis (FIA). METHODS: We analyzed data from the European Anaphylaxis Registry applying an age- and sex-matched analysis of associations (Cramer's V) for single food triggers and calculated odds ratios (ORs) for severe FIA. RESULTS: We identified 3,427 cases of confirmed FIA showing an age-dependent elicitor ranking (for children: peanut, cow's milk, cashew, and hen's egg; and for adults: wheat flour, shellfish, hazelnut, and soy). The age- and sex-matched analysis revealed defined symptom patterns for wheat and cashew. Wheat-induced anaphylaxis was more frequently associated with cardiovascular symptoms (75.7%; Cramer's V = 0.28) and cashew-induced anaphylaxis with gastrointestinal symptoms (73.9%; Cramer's V = 0.20). Furthermore, concomitant atopic dermatitis was slightly associated with anaphylaxis to hen's egg (Cramer's V = 0.19) and exercise was strongly associated with anaphylaxis to wheat (Cramer's V = 0.56). Additional factors influencing the severity were alcohol intake in wheat anaphylaxis (OR = 3.23; CI, 1.31-8.83) and exercise in peanut anaphylaxis (OR = 1.78; CI, 1.09-2.95). CONCLUSIONS: Our data show that FIA is age-dependent. In adults, the range of elicitors inducing FIA is broader. For some elicitors, the severity of FIA seems to be related to the elicitor. These data require confirmation in future studies considering a clear differentiation between augmentation and risk factors in FIA.

Specific IgE against the house dust mite allergens Der p 5, 20 and 21 influences the phenotype and severity of atopic diseases.

BACKGROUND: House dust mites (HDM) are among the most important sources for airborne allergens with high relevance for atopic diseases. Routine tests contain only 4 of 32 registered allergens of Dermatophagoides pteronyssinus. Clinical relevance and pathomechanistic properties of many allergens are not well understood. OBJECTIVE: The association of several HDM allergens with allergic rhinitis, allergic asthma, and atopic dermatitis was investigated to identify allergens with biomarker potential and to transfer them into diagnostics. METHODS: Eight out of nine D. pteronyssinus allergens (nDer p 1, rDer p 2, rDer p 5, rDer p 7, rDer p 10, rDer p 13, rDer p 20, rDer p 21, rDer p 23) were recombinantly expressed and purified. Sensitization patterns of 384 HDM-allergic individuals exhibiting different clinical phenotypes were analyzed with a serum-saving multiplex array. RESULTS: Sensitization to more than three mite allergens (sensitization count) was associated with allergic asthma and/or atopic dermatitis. Reactions to Der p 5 and Der p 21 were more frequent in allergic asthma compared to allergic rhinitis. Atopic dermatitis patients were more often sensitized to Der p 5, Der p 20, and Der p 21 among others. Der p 20-IgE > 80 kU/L was associated with severe atopic dermatitis in 75% of patients. CONCLUSION: This study demonstrates the clinical importance of the sensitization count and of certain allergens (Der p 5, Der p 20, and Der p 21) not available for routine diagnostics yet. Implementing them as well as the sensitization count in diagnostic measures will improve diagnosis and risk assessment of HDM-allergic patients.

Breastfeeding: Maternally Transferred Allergens in Breast Milk: Protective or Sensitizing?

According to a thorough literature search, the following allergen sources have been associated with allergy symptoms in the exclusively breastfed child: hen's egg, cow's milk, peanut, trout. Subsequently, several studies use the advantage of molecular allergology and investigate the potential transfer of single allergens into breastmilk. This is shown for caseins, whey proteins, gliadin, ovalbumin, ovomucoid, the peanut allergens Ara h 2 and Ara h 6, as well as the inhalant allergens Der p 1 and Blo t 5. It is still a matter of debate whether or not food allergens transferred via breastfeeding to the baby promote allergic sensitization or induce tolerance and via which mechanisms they may shift the immune response to the one or other side. Noteworthy, some breastfed children are described to be sensitized to foods before being exposed to solid foods, and this exposure may have occurred through breastmilk. In the light of these findings the investigation of food allergens transferred from the mother's diet into breastmilk and their impact on sensitization or allergy prevention remains a current topic in research. This review describes breastmilk in its composition and provides data on the identification of food allergens therein including human and mouse studies.

Update of the S2k guideline on the management of IgE-mediated food allergies.

Not available.

Effect of comorbid pulmonary disease on the severity of COVID-19: A systematic review and meta-analysis.

Coronavirus disease 2019 (COVID-19) caused by infection with severe acute respiratory syndrome coronavirus 2 was first detected in Wuhan, China, in late 2019 and continues to spread worldwide. Persistent questions remain about the relationship between the severity of COVID-19 and comorbid diseases, as well as other chronic pulmonary conditions. In this systematic review and meta-analysis, we aimed to examine in detail whether the underlying chronic obstructive pulmonary diseases (COPD), asthma and chronic respiratory diseases (CRDs) were associated with an increased risk of more severe COVID-19. A comprehensive literature search was performed using five international search engines. In the initial search, 722 articles were identified. After eliminating duplicate records and further consideration of eligibility criteria, 53 studies with 658,073 patients were included in the final analysis. COPD was present in 5.2% (2191/42,373) of patients with severe COVID-19 and in 1.4% (4203/306,151) of patients with non-severe COVID-19 (random-effects model; OR = 2.58, 95% CI = 1.99-3.34, Z = 7.15, p < 0.001). CRD was present in 8.6% (3780/44,041) of patients with severe COVID-19 and in 5.7% (16,057/280,447) of patients with non-severe COVID-19 (random-effects model; OR = 2.14, 95% CI = 1.74-2.64, Z = 7.1, p < 0.001). Asthma was present in 2.3% (1873/81,319) of patients with severe COVID-19 and in 2.2% (11,796/538,737) of patients with non-severe COVID-19 (random-effects model; OR = 1.13, 95% CI = 0.79-1.60, Z = 0.66, p = 0.50). In conclusion, comorbid COPD and CRD were clearly associated with a higher severity of COVID-19; however, no association between asthma and severe COVID-19 was identified.

Innovative robust basophil activation test using a novel gating strategy reliably diagnosing allergy with full automation.

BACKGROUND: Allergy is one of the most common chronic diseases in Europe. Therefore, an increased need for specific and sensitive diagnostic tests that truly detect allergy exists. This study aimed at establishing a highly specific high-throughput and automated basophil activation test (BAT) that proves the existence of an allergy with utmost probability. METHODS: BAT from 1104 samples was analyzed; a novel gating strategy with three antibodies (FcεRIα, CD203c, CD63) was established and compared with our published protocol (12 antibodies). Based on the novel gating strategy, storage conditions, automated measurement, and analyses using R (1376 samples out of 1389) were optimized to set up a high-throughput BAT. RESULTS: No differences in sensitivity and specificity were found between the novel three antibody (FcεRIα, CD203c, CD63) and the 12 antibody gating strategy or between automated and manually analyzed samples (saving up to 90% of labor time). The time frame for basophil activation measurement after blood donation has been extended considerably (whole blood storage ≤7 days (RT) and 17 days (4°C) prior to BAT preparation and measurement). Respective storage conditions were optimized for samples after stimulation, staining, and preparation (≤7 days (RT) and 28 days (4°C)). These achievements were confirmed by a nationwide ring trial showing robustness and applicability of our BAT on a variety of flow cytometers. CONCLUSION: Our considerable optimizations overcame the hurdles that until now prevented the BAT from being used as high-throughput allergy diagnostic test in routine laboratories and shall allow for collaborative studies between clinics and research centers.

Wheat Anaphylaxis in Adults Differs from Reactions to Other Types of Food.

BACKGROUND: Wheat is one of the most commonly consumed foods and a known elicitor of anaphylaxis in children and adults. Reactions in adults are often cofactor dependent and characterized by a prolonged time between food intake and the onset of symptoms making the diagnosis of wheat anaphylaxis challenging. OBJECTIVE: To characterize a cohort of patients with the history of wheat anaphylaxis to better understand this atypical phenotype of anaphylaxis. METHODS: Data from the European Anaphylaxis Registry from 2007 to 2019 (n = 10,636) including 250 patients (213 adults and 37 children) with a history of anaphylaxis caused by wheat were analyzed. RESULTS: Wheat was the most common food elicitor of anaphylaxis in adults in the registry in Central Europe. Reactions to wheat in adults were frequently associated with exercise as a cofactor (82.8%) and partially delayed (57.5%). Only 36.9% of patients had atopic comorbidities, which was uncommonly low for adult patients allergic to other kinds of foods (63.2%). Anaphylaxis to wheat presented frequently with cardiovascular symptoms (86.7%) including severe symptoms such as loss of consciousness (41%) and less often with respiratory symptoms (53.6%). The reactions to wheat were more severe than reactions to other foods (odds ratio [OR] = 4.33), venom (OR = 1.58), or drugs (OR = 2.11). CONCLUSIONS: Wheat is a relevant elicitor of anaphylaxis in adults in Central Europe. Wheat anaphylaxis is highly dependent on the presence of cofactors and less frequently associated with atopic diseases compared with other food allergies. More data on mechanisms of wheat-induced anaphylaxis are required to develop preventive measures for this potentially life-threatening disease.

Omalizumab ensures compatibility to bee venom immunotherapy (VIT) after VIT-induced anaphylaxis in a patient with systemic mastocytosis.

BACKGROUND: Systemic reactions and anaphylaxis due to Hymenoptera venoms occur in up to 7.5% of the European population. Fatal sting reactions are very rare. Serum tryptase levels should be measured in all patients with a history of severe reactions in order to detect mastocytosis and to determine the risk of severe reactions to venom immunotherapy (VIT). The risk to experience severe or even fatal anaphylaxis due to insect stings is quite high in patients with mastocytosis. Therefore, lifelong VIT is recommended in these highly threatened patients. Multicenter studies involving a large population report that up to 20% of patients undergoing VIT have intolerance and systemic reactions to immunotherapy. Some of these side effects occur repeatedly and cannot be managed by standard treatment. A pre-treatment with the anti-IgE antibody omalizumab was useful in many cases. However, omalizumab is not approved for the indication anaphylaxis. Therefore, there is still no defined protocol for omalizumab pre-treatment, and the optimal duration, dosage as well as long-time benefits are still unclear. CASE REPORT: We present a 60-year-old female patient with mastocytosis who developed a severe anaphylactic reaction during initiation of bee VIT. Serum tryptase was elevated, and a KIT mutation D816V was subsequently confirmed. Component-resolved diagnostic tests revealed specific IgE antibodies to recombinant Api m 1 only. The patient was treated with 150 mg omalizumab, administered subcutaneously 5 weeks, 3 weeks, and 1 week prior to re-start of immunotherapy and for 2 months in parallel to VIT. Updosing was done by a 7-day rush schedule. During this period, no anaphylactic reaction developed, and the bee VIT was well tolerated with up to 200 µg bee venom. The patient is currently in the 3rd year of treatment and tolerates the treatment very well. CONCLUSION: Omalizumab may be used as a premedication in patients with mastocytosis who do not tolerate VIT. Although there is no consensus on the treatment protocol, treatment for 2 - 6 months is considered adequate. The long-term benefits of such treatment require further research.

Carbohydrate epitopes currently recognized as targets for IgE antibodies.

Until recently, glycan epitopes have not been documented by the WHO/IUIS Allergen Nomenclature Sub-Committee. This was in part due to scarce or incomplete information on these oligosaccharides, but also due to the widely held opinion that IgE to these epitopes had little or no relevance to allergic symptoms. Most IgE-binding glycans recognized up to 2008 were considered to be "classical" cross-reactive carbohydrate determinants (CCD) that occur in insects, some helminths and throughout the plant kingdom. Since 2008, the prevailing opinion on lack of clinical relevance of IgE-binding glycans has been subject to a reevaluation. This was because IgE specific for the mammalian disaccharide galactose-alpha-1,3-galactose (alpha-gal) was identified as a cause of delayed anaphylaxis to mammalian meat in the United States, an observation that has been confirmed by allergists in many parts of the world. Several experimental studies have shown that oligosaccharides with one or more terminal alpha-gal epitopes can be attached as a hapten to many different mammalian proteins or lipids. The classical CCDs also behave like haptens since they can be expressed on proteins from multiple species. This is the explanation for extensive in vitro cross-reactivity related to CCDs. Because of these developments, the Allergen Nomenclature Sub-Committee recently decided to include glycans as potentially allergenic epitopes in an adjunct section of its website (www.allergen.org). In this article, the features of the main glycan groups known to be involved in IgE recognition are revisited, and their characteristic structural, functional, and clinical features are discussed.

Identification and Purification of Novel Low-Molecular-Weight Lupine Allergens as Components for Personalized Diagnostics.

Lupine flour is a valuable food due to its favorable nutritional properties. In spite of its allergenic potential, its use is increasing. Three lupine species, Lupinus angustifolius, L. luteus, and L. albus are relevant for human nutrition. The aim of this study is to clarify whether the species differ with regard to their allergen composition and whether anaphylaxis marker allergens could be identified in lupine. Patients with the following characteristics were included: lupine allergy, suspected lupine allergy, lupine sensitization only, and peanut allergy. Lupine sensitization was detected via CAP-FEIA (ImmunoCAP) and skin prick test. Protein, DNA and expressed sequence tag (EST) databases were queried for lupine proteins homologous to already known legume allergens. Different extraction methods applied on seeds from all species were examined by SDS-PAGE and screened by immunoblotting for IgE-binding proteins. The extracts underwent different and successive chromatography methods. Low-molecular-weight components were purified and investigated for IgE-reactivity. Proteomics revealed a molecular diversity of the three species, which was confirmed when investigated for IgE-reactivity. Three new allergens, L. albus profilin, L. angustifolius and L. luteus lipid transfer protein (LTP), were identified. LTP as a potential marker allergen for severity is a valuable additional candidate for molecular allergy diagnostic tests.

Biologics for atopic diseases: Indication, side effect management, and new developments.

With the advent of biologicals, more and more therapeutics are available that specifically address specific switch points in the pathomechanism of immunologically dominated diseases. Thus, the focus of diagnostics and therapy (precision medicine) is more on the individual disease characteristics of the individual patient. Regarding the different phenotypes of atopic diseases, severe asthma was the first entity for which biologicals were approved, followed by urticaria, and finally atopic dermatitis and chronic rhinosinusitis with nasal polyps. Experience in the treatment of severe bronchial asthma has shown that the intensity of the response to biological therapy depends on the quality of clinical and immunological phenotyping of the patients. This also applies to different diseases of the atopic form, as patients can suffer from several atopic diseases at the same time, each with different characteristics. Biologics are already emerging that may represent a suitable therapy for allergic bronchial asthma, which often occurs together with severe neurodermatitis, and chronic rhinosinusitis with nasal polyps. In practice, however, the question of possible combinations of biologicals for the therapy of complex clinical pictures of individual patients is increasingly arising. In doing so, the side effect profile must be taken into account, including hypersensitivity reactions, whose diagnostic and logistical management must aim at a safe and efficient therapy of the underlying disease. Increased attention must also be paid to biological therapy in pregnancy and planned (predictable) vaccinations as well as existing infections, such as SARS-CoV-2 infection. Before starting a biological therapy, the immune status should be checked with regard to chronic viral and bacterial infections and, if necessary, the vaccination status should be refreshed or missing vaccinations should be made up for before starting therapy. Currently, reliable data on the effect of biologicals on the immunological situation of SARS-CoV-2 infection and COVID-19 are not available. Therefore, research and development of suitable diagnostic methods for detection of immunologically caused side effects as well as detection of potential therapy responders and non-responders is of great importance.

Combination of immunotherapies for severe allergic asthma.

INTRODUCTION: Difficult-to-treat or severe persistent asthma accounts for 5-10% of the asthma population worldwide. However, this group of patients creates a higher burden on health systems due to their morbidity and need for long-term and additional treatment. Biological drugs constitute an alternative therapy in the treatment of patients with refractory asthma. In cases where inhalant allergy is part of the pathomechanism, allergen-specific immunotherapy (SIT) is a causative treatment option for allergic asthma. However, SIT is contraindicated for uncontrolled asthma and cannot be administered according to the guidelines. This is due to the risk of further worsening of uncontrolled asthma during treatment. CASE STUDY: We herein report a case of a 67-year-old male with severe allergic asthma who was successfully treated with SIT after asthma control was achieved by using target treatments. RESULTS: Complete control of asthma was achieved, and SIT with allergens from early flowering trees (birch-alder-hazel) was administered. Further, no asthmatic exacerbations or decrease in respiratory function occurred during the 15 months of treatment with mepoluzimab. He did not need any oral glucocorticosteroids. CONCLUSION: The case report presented here suggests the effectiveness of an individualized approach and phenotype-specific treatment of patients who cannot receive allergen-specific immunotherapy due to the contraindication uncontrolled asthma and who receive SIT after asthma control is achieved by using target treatments.

IgE Epitope Profiling for Allergy Diagnosis and Therapy - Parallel Analysis of a Multitude of Potential Linear Epitopes Using a High Throughput Screening Platform.

Immunoglobulin E (IgE) is pivotal for manifestation and persistence of most immediate-type allergies and some asthma phenotypes. Consequently, IgE represents a crucial target for both, diagnostic purposes as well as therapeutic approaches. In fact, allergen-specific immunotherapy - aiming to re-route an IgE-based inflammatory response into an innocuous immune reaction against the allergen - is the only curative approach for IgE-mediated allergic diseases known so far. However, this requires the cognate allergen to be known. Unfortunately, even in well-characterized allergics or asthmatics, often just a small fraction of total IgE can be assigned to specific target allergens. To overcome this knowledge gap, we have devised an analytical platform for unbiased IgE target epitope detection. The system relies on chemically produced random peptide libraries immobilized on polystyrene beads ("one-bead-one-compound (OBOC) libraries") capable to present millions of different peptide motifs simultaneously to immunoglobulins from biological samples. Beads binding IgE are highlighted with a fluorophore-labeled anti-IgE antibody allowing fluorescence-based detection and isolation of positives, which then can be characterized by peptide sequencing. Setting-up this platform required an elaborate optimization process including proper choice of background suppressants, secondary antibody and fluorophore label as well as incubation conditions. For optimal performance our procedure involves a sophisticated pre-adsorption step to eliminate beads that react nonspecifically with anti-IgE secondary antibodies. This step turned out to be important for minimizing detection of "false positive" motifs that otherwise would erroneously be classified as IgE epitopes. In validation studies we were able to retrieve artificial test-peptide beads spiked into our library by using IgE directed against those test-peptides at physiological concentrations (≤20 IU/ml of specific IgE), and disease-relevant bead-bound epitopes of the major peanut allergen Ara h 2 by screening with sera from peanut allergics. Thus, we established a platform with which one can find and validate new immunoglobulin targets using patient material which displays a largely unknown immunoglobulin repertoire.