Characterisation of major and minor Dermatophagoides allergens in canine atopic dermatitis

Atopic dermatitis is a well-recognised chronic inflammatory skin disease of humans and dogs. Most atopic dogs are sensitised to Dermatophagoides mites. The aim of this study was to characterise allergens in different Dermatophagoides species using polyclonal and monoclonal antibodies to canine IgE. Western blots were prepared from crude extracts of D farinae, D pteronyssinus and D microceras, and purified group 1 and 2 allergens under reducing and non-reducing conditions. They were probed with sera from atopic (n = 33) and healthy (n = 27) dogs. There was no significant difference in the sensitivity or specificity between the polyclonal and monoclonal sera in detecting Dermatophagoides -specific IgE. Major allergens common to both D farinae and D pteronyssinus were detected at 97-98 kDa, 103-104 kDa and 134-139 kDa on both reducing and non-reducing blots. Major allergens at 84-85 kDa, 65-69 kDa and 44-45 kDa were only recognised on reducing blots, suggesting that these are fragments of the larger allergens. Only a few sera recognised group 1 or 2 allergens on blots of crude extracts or purified allergens. These results confirm that, in atopic dogs, high molecular weight allergens are the most important Dermatophagoides allergens, rather than the low molecular weight group 1 and 2 proteins.     

Characterization and cloning of a major high molecular weight house dust mite allergen (Der f 15) for dogs

Although house dust mites (HDM(s)) are important elicitors of canine allergy, the low molecular weight molecules defined as major allergens for humans do not appear to be major allergens for dogs. Western blotting of Dermatophagoides farinae (D. farinae) extracts with sera from sensitized dogs showed that the majority of animals had IgE antibodies specific for two proteins of apparent molecular weights of 98 and 109kDa (98/109kDa). The N-terminal sequences of these two proteins were identical, suggesting they were very closely related, and sequencing of internal peptides showed the protein(s) to have homology with insect chitinases. A purified preparation of 98/109kDa proteins elicited positive intradermal skin tests (IDST(s)) in a group of well-characterized atopic dogs sensitized to D. farinae, but not in normal dogs. A rabbit polyclonal antiserum raised against the purified proteins was used to immunoscreen a D. farinae cDNA library. The mature coding region of the isolated chitinase cDNA predicts a protein of 63.2kDa; sequence analysis and glycan detection blotting suggest that the molecule is extensively O-glycosylated. Monoclonal antibodies made against the purified native protein were used to localize the chitinase in sections of whole D. farinae mites. The protein displayed an intracellular distribution in the proventriculus and intestine of the mite, suggesting that it has a digestive, rather than a moulting-related, function. The high prevalence of IgE antibodies to this antigen in canine atopic dermatitis makes it a major HDM allergen for dogs, and the protein has been formally designated Der f 15.     

Quantitation of house dust mites and house dust mite allergens in the microenvironment of dogs

OBJECTIVE: To quantitate the density of Dermatophagoides farinae and D pteronyssinus and concentrations of house dust mite (HDM) allergens (Der f 1, Der p 1, and Group 2 allergens) in the indoor microenvironment of dogs. SAMPLE POPULATION: 50 homes in Columbus, Ohio. PROCEDURES: In each home, samples of dust were collected from 3 locations in which dogs spent most time. Whenever possible, the species of mites collected was identified. Mite density (mites/g of dust) was assessed, and allergen concentrations were assayed by standardized ELISAs. Relative humidity and temperature in each home were monitored during a 5-day period. Characteristics of homes and sample sources were evaluated. RESULTS: Dust samples from all 50 homes contained > or = 1 HDM allergen; Der f 1 and Der p 1 were detected in 100 and 74% of homes, respectively. Fifteen homes had HDMs; compared with D pteronyssinus, D farinae was found more commonly (14/15 homes) and at a higher density. Basements, homes without central air-conditioning, and dog beds that were > or = 1 year old had high HDM allergen concentrations. Homes with > or = 2 microg of Der f 1 or Group 2 allergens/g of dust or > or = 100 mites/g of dust were significantly more likely to have a maximum relative humidity > or = 75%. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated the presence of HDMs and HDM allergens in the specific microenvironment of dogs in homes. Factors associated with high levels of exposure were identified, which may be associated with increased risk for sensitization and development of atopic diseases.     

Determination of antigenic proteins of housedust mites in 90 dogs suffering from atopic dermatitis

Housedust mites, Dermatophagoides pteronyssinus (D. pteronyssinus) and Dermatophagoides farinae (D. farinae), are the important causative agents of allergic diseases in human and animals. By using 165 dogs suffering from atopic dermatitis (AD), serum levels of immunogloblin E (IgE) antibody against 25 kinds of allergen including housedust mites were determined. Housedust mites were the most frequent allergen against which 90 of the 165 allergic dogs (54.5%) by IMMUNODOT assay. With the further analysis of immunoblotting assay in the 90 dogs sensitized with housedust mites, antigenic proteins of housedust mites recognized by IgE antibodies were with the apparent molecular masses of 15, 76, 90, 98, and 170-kD. Among them, the 15-kD protein that might be identical to Group 2 antigens (Der f2, Der p2) was prominently observed (52/90). This study indicates that about a half of dogs with AD were sensitized to housedust mites, suggesting that Group 2 antigens of housedust mites may be a major allergen in canine AD.     

Quantitation of house dust mites and house dust mite allergens in the microenvironment of dogs

OBJECTIVE: To quantitate the density of Dermatophagoides farinae and D pteronyssinus and concentrations of house dust mite (HDM) allergens (Der f 1, Der p 1, and Group 2 allergens) in the indoor microenvironment of dogs. Sample Population-50 homes in Columbus, Ohio. PROCEDURES: n each home, samples of dust were collected from 3 locations in which dogs spent most time. Whenever possible, the species of mites collected was identified. Mite density (mites/g of dust) was assessed, and allergen concentrations were assayed by standardized ELISAs. Relative humidity and temperature in each home were monitored during a 5-day period. Characteristics of homes and sample sources were evaluated. RESULTS: Dust samples from all 50 homes contained > or = 1 HDM allergen; Der f 1 and Der p 1 were detected in 100 and 74% of homes, respectively. Fifteen homes had HDMs; compared with D pteronyssinus, D farinae was found more commonly (14/15 homes) and at a higher density. Basements, homes without central air-conditioning, and dog beds that were > or = 1 year old had high HDM allergen concentrations. Homes with > or = 2 microg of Der f 1 or Group 2 allergens/g of dust or > or = 100 mites/g of dust were significantly more likely to have a maximum relative humidity > or = 75%. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate the presence of HDMs and HDM allergens in the specific microenvironment of dogs in homes. Factors associated with high levels of exposure were identified, which may be associated with increased risk for sensitization and development of atopic diseases.     

Characterization of house dust mite and scabies mite allergens by use of canine serum antibodies

OBJECTIVE: To identify the major allergenic proteins from the 3 main species of dust mites to which dogs react (Dermatophagoides farinae, D. pteronyssinus, and Euroglyphus maynei) and evaluate the potential cross-reactivity of dust mite allergens with antigens from the ectoparasitic mite Sarcoptes scabiei var canis. SAMPLE POPULATION: Sera from 83 dogs with atopic dermatitis. PROCEDURE: Sodium dodecylsulfate-polyacrylamide gel electrophoresis and immunoblotting using serum from atopic dogs was used to identify IgE-binding proteins in extracts of the 4 mite species. RESULTS: Sera of atopic dogs contained IgE against 23, 17, 25, and 17 allergens from D. farinae, D. pteronyssinus, E. maynei, and S. scabiei, respectively. Unlike the situation for humans, the major allergens for dogs are mostly proteins that are larger than 90 kd molecular weight. Dermatophagoides farinae and E. maynei appear to be more allergenic for dogs than is D. pteronyssinus. Some dogs with serum IgE against dust mites also had IgE against antigens of S. scabiei var canis. CONCLUSIONS AND CLINICAL RELEVANCE: Multiple dust mite allergens induce an IgE response in dogs. These allergens are mostly greater than 90 kd molecular weight.     

Prevalence of house dust mites and dermatophagoides group 1 antigens collected from bedding, skin and hair coat of dogs in south-west England

The house dust mites Dermatophagoides farinae (Df) and D. pteronyssinus (Dpt) are commonly implicated as allergens causing canine atopic dermatitis in the UK. However, there are few studies that characterize the exposure of UK pet dogs to these mites. The objectives of this study were to determine the prevalence of the mite species on the skin, hair coat and bedding of a population of pet dogs. Dust samples (n = 68) were collected from both dogs and their beds using a standardized vacuuming technique and stored at -20 degrees C. Mites were identified using accepted morphological criteria. House dust mite allergen concentrations were assayed using standardized ELISA for Dpt and Df group 1 allergens (Der p 1 and Der f 1). Mites were identified in 15/68 samples (22%) and Dpt was the most common. Df mites were not present. Der p 1 allergens were detected in 60% of samples, and Der f 1 in 6% of samples. There were no significant differences between the number of Der p 1 positive samples from dogs and the number of those from their bedding, or between the average Der p 1 concentrations from dogs and the number of those from their bedding. Contrary to studies elsewhere in Europe and the USA, these findings support studies of human asthma patients in the UK, where exposure to Df is rare, but to Dpt is common. As the prevalence of positive intradermal and serological reactions to Df in atopic dogs is high, further investigations are warranted to clarify true Df hypersensitivity or potential immunological cross-reactivity between mite allergens.     

Peripheral blood mononuclear cell responses to Dermatophagoides farinae in canine atopic dermatitis

Atopic dermatitis is a chronic inflammatory and pruritic skin disease commonly seen in dogs and humans that is characterised by the presence of allergen-specific IgE. Data from skin tests and serological analysis suggest that the house dust mite Dermatophagoides farinae is the most important allergen in dogs with atopic dermatitis. The aim of this study was to determine if D. farinae specific peripheral blood mononuclear cell (PBMC) responses could be detected in dogs with atopic dermatitis. PBMCs were isolated by the density centrifugation from dogs with atopic dermatitis that were skin test positive for D. farinae, dogs with atopic dermatitis that were skin test negative for D. farinae, and healthy dogs. Cells were cultured with increasing concentrations of the D. farinae extract, no antigen, vaccine antigens or concanavalin A (ConA). There was significantly greater responsiveness of PBMCs from the D. farinae positive dogs than from either the D. farinae negative or healthy dogs (ANOVA, P<0.05). In contrast, no significant differences were observed in the control responses between the three groups. This is the first study to demonstrate that D. farinae specific circulating memory cells are involved in the pathogenesis of canine house dust mite hypersensitivity.     

Prevalence of the group 1 Dermatophagoides allergens Der p 1 and Der f 1 in homes with no dogs, healthy dogs and Dermatophagoides-sensitized atopic dogs in Liverpool

Dermatophagoides farinae is a frequent allergen in canine atopic dermatitis despite its reported scarcity in the UK, and the aim of this study was to determine whether dogs were uniquely exposed to this species. Der f 1 and Der p 1 in dust collected from living room carpets, bedroom carpets and dog beds of 13 houses with no dogs, 13 with healthy dogs, and 16 with Dermatophagoides-sensitized atopic dogs were quantified by ELISA. Der p 1 levels (microg g(-1) house dust) were significantly higher than Der f 1 in living rooms (Der p 1 median = 1.9, 95% CI = 2.05-6.32, n = 42; Der f 1 median = 0.07, 95% CI = 0.01-0.06, n = 42), bedrooms (Der p 1 median = 4.35, SD = 5.52; Der f 1 median = 0.01, 95% CI = 0.001-0.1, n = 42) and dog beds (Der p 1 median = 1.04, 95% CI = 1.4-8.1, n = 29; Der f 1 median = 0.008, 95% CI = 0.01-0.04, n = 29) (P < 0.0001). Living rooms in houses without dogs had significantly greater Der p 1 levels (median = 7.0, 95% CI = 3.53-15.8, n = 13) than houses with healthy (median = 1.19, 95% CI = 0.44-3.49, n = 13) or atopic dogs (median = 0.78, 95% CI = 0.63-2.42, n = 16) (P = 0.0004). Environmental flea control in living rooms and washing dog beds was associated with significantly reduced Der p 1 levels. This confirms that D. pteronyssinus is common but D. farinae is rare in the sampling area. Apparent sensitization to D. farinae is probably due to cross-reaction. A combination of environmental measures could reduce allergen exposure.     

Sensitivity patterns to house dust mites and forage mites in atopic dogs: 150 cases

This study investigated intradermal test reactions to extracts of six species of mites in 150 dogs with atopic dermatitis. At least one positive reaction was seen in 120 animals (80%). Dermatophagoides farinae attracted the highest number of positive reactions (108 dogs, 90% of dogs and 72% of atopic dogs showing positive reactions). Positive reactions to other mites were not uncommon, with many dogs testing positive for Dermatophagoides pteronyssinus (32% of dogs tested), Acarus siro (35%), Tyrophagus putrescentiae (30%), Glycyphagus domesticus (27%) and Lepidoglyphus destructor (23%). Sensitivity to D. farinae alone occurred commonly (57% of cases), but multiple sensitivities were seen frequently with the other mites. Cases of sensitivity to only one mite were also seen: D. pteronyssinus (five cases), T. putrescentiae (one case) and G. domesticus (one case). Further studies are needed to appreciate more clearly the precise role played by the different species of mite in canine atopic dermatitis.     

Importance of house dust and storage mites in canine atopic dermatitis in the geographic region of Galicia, Spain

Sensitisation to mites is frequent in atopic dogs. The main mite genus involved in canine atopic dermatitis is Dermatophagoides. The importance of storage mite allergens in dogs has been controversial. The aim of this study was to evaluate the sensitisation rates against storage mites (Lepidoglyphus destructor and Tyrophagus putrescentiae) and house dust mites (Dermatophagoides farinae and D. pteronyssinus) in atopic dogs from Galicia, a highly humid and temperate region of Spain, using a FcepsilonRIalpha-based immunoglobulin E (IgE) in vitro test. The study was performed on 95 dogs suffering from atopic dermatitis and presenting detectable specific serum IgE levels: 91.6% of the dogs tested positive for storage mites, whereas sensitisation to house dust mites was detected in 87.4%. These results indicate the importance of storage mites in this specific geographic area.     

Production of GM-CSF Mediated by Cysteine Protease of Der f in Canine Keratinocytes

House dust mite (HDM) allergens are the most common allergens for induction of IgE-mediated hypersensitivity. Recently, epicutaneous sensitization with HDM allergens has been emphasized in the development of atopic dermatitis (AD) by producing various soluble factors in keratinocytes. Among the soluble factors, GM-CSF is a key molecule that activates Langerhans cells, antigen-presenting cells in the epidermis. In the present study, we investigated the effects of Dermatophagoides farinae (Der f) on GM-CSF production in a canine keratinocyte cell line, CPEK. CPEKs were found to produce GM-CSF upon stimulation by Der f. The GM-CSF production was suppressed by addition of a cysteine protease inhibitor. The present results suggest that cysteine protease-derived Der f may be an initiator of allergic inflammation by inducing the production of GM-CSF in keratinocytes.     

Reactivity to intradermal injection of extracts of Dermatophagoides farinae, Dermatophagoides pteronyssinus, house dust mite mix, and house dust in dogs suspected to have atopic dermatitis: 115 cases (1996-1998)

OBJECTIVE: To compare reactivities to intradermal injection of extracts of Dermatophagoides farinae, Dermatophagoides pteronyssinus, house dust mite mix, and house dust in dogs suspected to have atopic dermatitis. DESIGN: Retrospective study. ANIMALS: 115 dogs. PROCEDURES: Records of all dogs suspected to have atopic dermatitis that underwent intradermal testing between October 1996 and July 1998 were reviewed. Reactivities to intradermal injection of crude mixed house dust mite (1:25,000 wt/vol) and crude house dust (25 PNU/ml) extracts were compared with reactivities to intradermal injection of individual extracts of D farinae and D pteronyssinus (1:50,000 wt/vol). RESULTS: Ninety dogs were confirmed to have atopic dermatitis including 61 of the 69 dogs with positive reactions to either or both of the individual house dust mite extracts. Intradermal testing with the mixed house dust mite extract had sensitivity of 75%, specificity of 96%, and accuracy of 83%. Intradermal testing with the house dust extract had sensitivity of 30%, specificity of 93%, and accuracy of 56%. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that use of crude mixed house dust mite and crude house dust extracts for intradermal testing in dogs is not as accurate a method of determining house dust mite hypersensitivity as is the use of individual D farinae and D pteronyssinus extracts mainly because of the high percentage of false-negative results. Extracts of individual house dust mites are recommended for intradermal testing of dogs suspected to have atopic dermatitis.     

Serum immunoglobulin E against storage mite allergens in dogs with atopic dermatitis

OBJECTIVE: To determine the prevalence of serum IgE against the storage mites Acarus siro, Blomia tropicalis, and Tyrophagus putrescentiae in a population of dogs with atopic dermatitis. SAMPLE POPULATION: Sera from 84 dogs with atopic dermatitis residing in various regions of the United States and Europe. PROCEDURE: Immunoblotting of sera from atopic dogs was used to identify proteins in mite extracts that bound IgE. RESULTS: 94% of the dogs had serum IgE against proteins in extracts of 1 or more of the storage mite species. Ninety-five, 92, and 89% of the storage mite-sensitive dogs had serum IgE against proteins in extracts of A siro, B tropicalis, and T putrescentiae, respectively. Eighty-two percent had serum IgE against at least 1 protein in all 3 species. Most of the major allergens had molecular weights > 80 kd. A greater percentage of the dog sera had IgE against storage mite proteins, compared with proteins of the house dust mites Dermatophagoides farinae and D pteronyssinus. CONCLUSION AND CLINICAL RELEVANCE: Many dogs with atopic dermatitis have serum IgE against many allergens of storage mites. Most of these allergens, like allergens of dust mites, had molecular weights > 80 kd. Storage mite sensitivity in dogs may be as important, if not more important, than dust mite sensitivity.     

Dermatophagoides farinae-specific IgG responses in atopic dogs undergoing allergen-specific immunotherapy with aqueous vaccines

The molecular and immunologic mechanisms associated with successful allergen-specific immunotherapy (ASIT) have not been completely elucidated. The aim of this study was to characterize the changes in Dermatophagoides farinae -specific IgG in atopic dogs undergoing ASIT using aqueous vaccines. Fifteen atopic dogs with a positive skin test reaction to D. farinae were treated with aqueous vaccines for a minimum of 2 months following a standard protocol. Serum samples were collected before and during therapy and used to probe Western blots containing separated proteins of D. farinae . IgG responses were detected using a polyclonal goat anticanine IgG antibody and a chromogenic substrate 3,3'-diaminobenzidine. The blots were analysed using a semiquantitative digital image analysis system that evaluated the number and molecular weight of bands, as well as their intensity, which was related to IgG concentration. Prior to ASIT, all dogs showed allergen-specific IgG responses to various antigens of D. farinae . During ASIT, there was a significant increase in the total quantity of D. farinae -specific IgG antibodies to various antigens from the mite ( P  = 0.015). Significant increases were observed for a 98-kDa band ( P  = 0.015), likely to be Der f 15; bands with molecular weights between 50 and 70 kDa ( P  = 0.012); and bands between 30 and 45 kDa ( P  = 0.035). These findings provide support for the hypothesis that ASIT induces IgG blocking antibodies to allergens known to be relevant in canine atopic dermatitis.     

Immunological findings in 3 dogs clinically diagnosed with allergic rhinitis

Three dogs clinically diagnosed with allergic rhinitis (AR) were examined for their immunological findings. House dust mites (HDM) such as Dermatophagoides farinae (DF) and D. pteronyssinus (DP) were identified as positive allergens in the 3 dogs with both intradermal skin test and serum antigen-specific IgE test. Lymphocyte blastogenic response of peripheral blood mononuclear cells (PBMCs) under stimulation with DF antigen in dogs with AR was higher than that in 4 healthy control dogs. Expression level of IL-4 mRNA in PBMCs obtained from the 3 AR dogs was higher than that in PBMCs obtained from 4 healthy control dogs before and after stimulation with DF antigen. Expression level of IFN-gamma mRNA in PBMCs was not different between the AR and control dogs before and after stimulation with DF antigen. These results suggested that allergic reaction to HDM antigen and T(H)2-type immune response were associated with the development of AR in 3 dogs examined in this study.     

Comparison of intradermal and serum testing for allergen-specific IgE using a Fcepsilon RIalpha-based assay in atopic dogs in the UK

Atopic dermatitis in dogs is a common allergic skin disease that affects substantial numbers of dogs in the UK. The purpose of this study was to compare the results of an intradermal test (IDT) and an in vitro test in a large cohort of dogs. Dogs were intradermal tested with Greer allergens (Greer Labs Inc, Lenoir, NC, USA) using standard techniques. At the same time blood samples were drawn and submitted for evaluation by ELISA using the ALLERCEPT Definitive Allergen Panels for allergen-specific IgE, a commercial assay that uses a biotinylated recombinant extracellular domain of the high affinity Fc-epsilon receptor alpha chain protein (Fcepsilon RIalpha). The allergens used in the two tests included grass, tree and weed pollens, moulds, flea saliva/whole flea extract and house dust mite species. The optical density readings from the ELISA for each allergen were compared with the results of the IDT for 265 dogs. The prevalence of positive reactions in the ELISA was equal to or greater than the results of the IDT in the case of almost all of the allergens, but two notable exceptions were the house dust mites Dermatophagoides farinae and Dermatophagoides pteronyssinus. These two allergens were the most common positive reactions by IDT (prevalence D. farinae 78.9%, D. pteronyssinus 66.4%). The results of the two tests were significantly different (McNemar's test, P<0.05) for 16 of the 22 allergens. The sensitivities of the ELISA compared to the IDT (where there were more than 3 dogs with positive reactions in both tests) varied between 19.3 and 77.1% (D. pteronyssinus 19.3% and D. farinae 67.9%) and the specificities varied between 64.2 and 96.6% (D. pteronyssinus 96.6% and D. farinae 89.3%).     

IgE sensitivity and cross-reactivity to crude and purified mite allergens (Der f 1, Der f 2, Der p 1, Der p 2) in atopic dogs sensitive to Dermatophagoides mite allergens

The present study investigates IgE-reactivity to crude and purified mite allergens by intradermal skin test (IDST), Immunodot method, and ELISA in atopic dogs sensitive to mite allergens, as well as the allergenic cross-reactivity between Dermatophgoides (D) farinae (DF) and D. pteronyssinus (DP) in dogs by IgE-ELISA inhibition. IDST and Immunodot method for crude mite allergens were performed for atopic dogs and 16 atopic dogs showed sensitivity to mite allergens. Of the 16 dogs, all dogs had anti-DF IgE and 11 had anti-DP IgE. We measured specific IgE to purified major allergens (Der f 1, Der f 2, Der p 1, Der p 2). Of the 16 atopic dogs, six had anti-Der f 1 IgE and seven had anti-Der f 2 IgE. Similarly, of the 16 dogs, six had anti-Der p 1 IgE and seven had anti-Der p 2 IgE. However, eight dogs had no specific IgE to these mite allergens. These dogs may be sensitive to other major mite allergens except Der 1 and Der 2. In the dogs that had both anti-DF and DP IgE, IgE binding to DF was greatly inhibited by DP, and reciprocal inhibition was observed. Based on these data, it appears that there is a strong cross-reactivity between DF and DP in dogs. Similarly, a cross-reactivity between DF and DP in purified allergens was also observed. IDST and Immunodot method are useful methods for the diagnosis of atopic diseases in dogs, and ELISA is a useful method for further investigation of IgE-reactivity for the allergens.     

High allergen-specific serum immunoglobulin E levels in nonatopic West Highland white terriers

Human and canine atopic dermatitis (AD) share an association with IgE specific to environmental allergens, but few studies have evaluated serum allergen‐specific IgE in nonatopic dogs. This study compared serum allergen‐specific IgE levels in 30 atopic and 18 nonatopic West Highland white terriers. Atopic dermatitis was confirmed using standard criteria. Nonatopic dogs were over 5 years of age and had no clinical signs or history of AD. Serum allergen‐specific IgE levels were measured with Allercept^® IgE ELISAs using a 48‐allergen Australian panel. Positive reactions were defined as ≥150 ELISA absorbance units. Intradermal tests were performed in 16 atopic dogs, either at the time of or at various times prior to serum collection. In atopic dogs, the most common positive ELISA and intradermal test results were to Dermatophagoides farinae (11 of 30 dogs), but there were no statistically significant correlations between results from the two methods for any allergen. In nonatopic dogs, multiple high‐positive ELISA reactions were reported to 45 of 48 allergens, most commonly D. farinae and Tyrophagus putrescentiae (17 of 18 dogs each). Positive ELISA results in nonatopic dogs were statistically significantly higher than those in atopic dogs for 44 of 48 allergens, including two allergens (D. farinae and Dermatophagoides pteronyssinus) commonly regarded as significant in canine AD. In conclusion, positive allergen‐specific IgE ELISAs were not specific for canine AD, and high allergen‐specific IgE levels were seen in nonatopic dogs. The clinical significance of this and whether it characterizes a protective phenotype is unclear.