Designing a Multimer Allergen for Diagnosis and Immunotherapy of Dog Allergic Patients

Nilsson, Ola; Neimert-Andersson, Theresa; Bronge, Mattias; Grundström, Jeanette; Sarma, Ranjana; Uchtenhagen, Hannes; Kikhney, Alexey; Sandalova, T.; Holmgren, Erik; Svergun, Dmitri I.; Achour, Adnane; Hage, Marianne van; Grönlund, Hans

doi:10.1371/journal.pone.0111041

Cited by 20 publications

(14 citation statements)

References 33 publications

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“…Typical symptoms of animal allergies are skin rashes, rhinitis, asthma, and occasionally anaphylaxis. It has been reported that serum levels of antigen-specific IgE in model mice subcutaneously sensitized to Can f 1 with Alum were significantly increased (14). Our results confirmed elevation of serum IgE and IL13 levels in mice sensitized with and without Alum by the same sensitization route in the same sensitization schedule; however, no typical animal allergy symptoms were observed.…”

Section: Il13 Production From Splenocytessupporting

confidence: 87%

“…Purified Can f 1 protein (endotoxin < 0.03 EU/μg; INDOOR Biotechnologies, VA, USA) and OVA (grade V; Sigma-Aldrich, MO, USA) were used as animal allergen and food allergen, respectively. Immunization of mice was performed as described previously (14). Briefly, 5 μg of each antigen with or without 1 mg of Alum (LSL, Tokyo, Japan) was dissolved in 100 μL of Dulbecco's phosphate buffered saline (PBS).…”

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confidence: 99%

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Comparison of immune response in mice sensitized to an animal allergen, Can f 1, and to a food allergen, ovalbumin

et al. 2019

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Can f 1 belongs to the lipocalin superfamily and is considered to be an animal allergen. The immune response induced by Can f 1 in mice was compared with that induced by ovalbumin (OVA), a typical food allergen. Female BALB/c and C57BL/6 mice (6 weeks of age) were subcutaneously injected with Can f 1 or OVA with or without aluminum hydroxide (Alum) three times with intervals of two weeks. Serum levels of total IgE or antigen-specific IgE and production of IL13 and IFNγ from splenocytes were analyzed. Immunization with Can f 1 or OVA increased serum levels of both total IgE and antigen-specific IgE significantly irrespective of Alum. These results indicate that Can f 1 and OVA were able to induce allergic sensitization in mice. Splenocyte production of IL13 in mice immunized with Can f 1 or OVA with and without Alum were significantly increased after stimulation with each antigen. However, IL13 levels in the mice immunized with Can f 1 with Alum were significantly lower than those immunized without Alum. Increases in IFNγ levels after stimulation with Can f 1 or OVA were not remarkable. No influence of genetic backgrounds of BALB/c and C57BL/6 mice was found. Although Can f 1 induced Th2 type immune responses as was also the case for immunization with OVA, an inhibitory effect of Alum on induction of IL13 was observed only in mice immunized with Can f 1. These results suggest that the immune mechanism for allergic sensitization with Can f 1 is different from that with OVA.An allergy is defined as a condition showing immunological hypersensitivity to a specific object. A food allergy is a type of allergy caused by ingestion of food. Ovalbumin (OVA) and casein derived from eggs and milk, respectively, are the most common causes of food allergies. An animal allergy is a hypersensitivity caused by animal substances such as dander, saliva, and urine and is characterized by skin rashes, rhinitis, asthma, and occasionally anaphylaxis. In the development of an animal allergy, exposure to allergens mostly occurs through inhala-tion or through skin contact. Animal models, especially mice, have significantly contributed to the elucidation of onset mechanisms of allergies and the development of diagnosis and therapy for allergies (7). OVA has been used most widely as an immunogen to establish allergy models in mice which develop asthma, atopic dermatitis, allergic rhinitis, anaphylaxis, and food allergy. However, studies of mouse models using animal allergens have been limited. Studies of mouse model sensitized to OVA have revealed that allergy is mediated by CD4 + T-helper type 2 (Th2) lymphocytes that, through the production of cytokines such as IL4, IL5 and IL13, orchestrate allergen-specific IgE synthesis and eventually eosinophilic inflammation in the target organs (16). Anaphylaxis can occur in OVA sensitized mice through two independent pathways; a pathway that

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Section: Il13 Production From Splenocytessupporting

confidence: 87%

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confidence: 99%

Comparison of immune response in mice sensitized to an animal allergen, Can f 1, and to a food allergen, ovalbumin

et al. 2019

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“…Nilsson and colleagues designed a tetrameric dog allergen molecule which comprised Can f 1, Can f 2, Can f 4, and Can f 6 [17••]. Allergens expressed in the E. coli tetramer were correctly folded, independently positioned, and retained their three-dimensional structure.…”

Section: Next-generation Innovationsmentioning

confidence: 99%

Technological Innovations for High-Throughput Approaches to In Vitro Allergy Diagnosis

Chapman

Wuenschmann

King

et al. 2015

Curr Allergy Asthma Rep

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Allergy diagnostics is being transformed by the advent of in vitro IgE testing using purified allergen molecules, combined with multiplex technology and biosensors, to deliver discriminating, sensitive, and high-throughput molecular diagnostics at the point of care. Essential elements of IgE molecular diagnostics are purified natural or recombinant allergens with defined purity and IgE reactivity, planar or bead-based multiplex systems to enable IgE to multiple allergens to be measured simultaneously, and, most recently, nanotechnology-based biosensors that facilitate rapid reaction rates and delivery of test results via mobile devices. Molecular diagnostics relies on measurement of IgE to purified allergens, the “active ingredients” of allergenic extracts. Typically, this involves measuring IgE to multiple allergens which is facilitated by multiplex technology and biosensors. The technology differentiates between clinically significant cross-reactive allergens (which could not be deduced by conventional IgE assays using allergenic extracts) and provides better diagnostic outcomes. Purified allergens are manufactured under good laboratory practice and validated using protein chemistry, mass spectrometry, and IgE antibody binding. Recently, multiple allergens (from dog) were expressed as a single molecule with high diagnostic efficacy. Challenges faced by molecular allergy diagnostic companies include generation of large panels of purified allergens with known diagnostic efficacy, access to flexible and robust array or sensor technology, and, importantly, access to well-defined serum panels form allergic patients for product development and validation. Innovations in IgE molecular diagnostics are rapidly being brought to market and will strengthen allergy testing at the point of care.

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“…In short, mouse antibodies directed to each dog allergen, Can f 1‐6, were added to ninety‐six–well MaxiSorp plates (Thermo Fisher Scientific) pre‐coated with goat‐anti mouse IgG (Jackson Immunoresearch, Ely, UK). The plates with allergen‐specific antibodies were incubated with equal amounts of sample and competitor, corresponding to in vivo biotinylated recombinant Can f 1‐2‐4‐6 for Can f 1, Can f 2, Can f 4, and Can f 6 measurements. In vitro biotinylated native Can f 3 and in vitro biotinylated recombinant Can f 5 was used for the Can f 3 and Can f 5 measurement, respectively.…”

Section: Methodsmentioning

confidence: 99%

Allergens in dog extracts: Implication for diagnosis and treatment

Wintersand

Asplund

Binnmyr

et al. 2019

Allergy

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Background: Five to ten percent of the population in affluent countries are allergic to dog. Diagnosis and treatment is based on allergen extracts from natural sources where composition and concentration are poorly defined.Objective: We aimed to quantify six dog allergens (Can f 1-6) in commercial skin prick test (SPT) solutions and to determine individual allergen profiles in dogs. Method:The allergen content of SPT solutions from five vendors and allergen source material from three anatomical sites were analyzed. Fur and saliva samples were collected from a mixed population of 120 dogs. Can f 1-6 were quantified by inhibition ELISA using purified recombinant or natural allergens and polyclonal or monoclonal antibodies. Allergenicity was analyzed by basophil activation test.

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Designing a Multimer Allergen for Diagnosis and Immunotherapy of Dog Allergic Patients

Cited by 20 publications

References 33 publications

<b>Comparison of immune response in mice sensitized to an animal allergen, Can f 1, and to a food allergen, </b><b>ovalbumin</b>

<b>Comparison of immune response in mice sensitized to an animal allergen, Can f 1, and to a food allergen, </b><b>ovalbumin</b>

Technological Innovations for High-Throughput Approaches to In Vitro Allergy Diagnosis

Allergens in dog extracts: Implication for diagnosis and treatment

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