Analysis of cytokine production by peanut-reactive T cells identifies residual Th2 effectors in highly allergic children who received peanut oral immunotherapy.

BACKGROUND: Only limited evidence is available regarding the cytokine repertoire of effector T cells associated with peanut allergy, and how these responses relate to IgE antibodies to peanut components. OBJECTIVE: To interrogate T cell effector cytokine populations induced by Ara h 1 and Ara h 2 among peanut allergic (PA) children in the context of IgE and to evaluate their modulation during oral immunotherapy (OIT). METHODS: Peanut-reactive effector T cells were analysed in conjunction with specific IgE profiles in PA children using intracellular staining and multiplex assay. Cytokine-expressing T cell subpopulations were visualized using SPICE. RESULTS: Ara h 2 dominated the antibody response to peanut as judged by prevalence and quantity among a cohort of children with IgE to peanut. High IgE (> 15 kU(A)/L) was almost exclusively associated with dual sensitization to Ara h 1 and Ara h 2 and was age independent. Among PA children, IL-4-biased responses to both major allergens were induced, regardless of whether IgE antibodies to Ara h 1 were present. Among subjects receiving OIT in whom high IgE was maintained, Th2 reactivity to peanut components persisted despite clinical desensitization and modulation of allergen-specific immune parameters including augmented specific IgG4 antibodies, Th1 skewing and enhanced IL-10. The complexity of cytokine-positive subpopulations within peanut-reactive IL-4(+) and IFN-γ(+) T cells was similar to that observed in those who received no OIT, but was modified with extended therapy. Nonetheless, high Foxp3 expression was a distinguishing feature of peanut-reactive IL-4(+) T cells irrespective of OIT, and a correlate of their ability to secrete type 2 cytokines. CONCLUSION: Although total numbers of peanut-reactive IL-4(+) and IFN-γ(+) T cells are modulated by OIT in highly allergic children, complex T cell populations with pathogenic potential persist in the presence of recognized immune markers of successful immunotherapy.

Infection with human rhinovirus 16 promotes enhanced IgE responsiveness in basophils of atopic asthmatics.

BACKGROUND: Rhinovirus and IgE act in concert to promote asthma exacerbations. While basophils are the principal cell type in the blood that is activated by IgE, their role in virus-induced asthma episodes remains elusive. OBJECTIVE: To monitor IgE responsiveness in circulating basophils of rhinovirus-infected atopic asthmatics during acute infection and convalescence. METHODS: The capacity for basophils to respond to IgE was assessed by testing the effects of allergen, or cross-linking anti-FcεRI and anti-IgE antibodies, on surface TSLP receptor in 24-hour PBMC cultures. Activation profiles of basophils from atopic asthmatics challenged intranasally with human rhinovirus 16 were monitored directly ex vivo or else in 24-hour cultures, at baseline (day 0), and then at days 4 and 21 post-challenge. RESULTS: Basophils in atopic asthmatics, but not in non-atopic controls, upregulated TSLP receptor upon IgE receptor ligation. The magnitude of this response was correlated with the proportion of serum total IgE that was allergen-specific (r = 0.615, P < 0.05). Following rhinovirus infection, all subjects developed nasal symptoms that peaked 3-5 days after viral challenge. Basophils displayed maximal IgE responsiveness 3 weeks post-challenge as judged by TSLP receptor levels in 24-hour cultures. No significant change in total IgE or specific IgE antibodies was detected during rhinovirus infection. By contrast, levels of IgE receptor-associated spleen tyrosine kinase, Syk, were increased on day 4 (P < 0.05), and elevated levels were also detected three weeks post-challenge. CONCLUSIONS AND CLINICAL RELEVANCE: Circulating basophils display increased IgE responsiveness 3 weeks after rhinovirus infection in atopic asthmatics. This observation, coupled with increased expression of Syk, implicates basophils in promoting, or else prolonging, rhinovirus-induced inflammation in atopic asthmatics.

Sensitization to food and inhalant allergens in relation to age and wheeze among children with atopic dermatitis.

BACKGROUND: Atopic dermatitis (AD) is common in children; however, persistence of AD with or without asthma is less common. Longitudinal studies remain limited in their ability to characterize how IgE antibody responses evolve in AD, and their relationship with asthma. OBJECTIVE: To use a cross-sectional study design of children with active AD to analyse age-related differences in IgE antibodies and relation to wheeze. METHODS: IgE antibodies to food and inhalant allergens were measured in children with active AD (5 months to 15 years of age, n = 66), with and without history of wheeze. RESULTS: Whereas IgE antibodies to foods persisted at a similar prevalence and titre throughout childhood, IgE antibodies to all aeroallergens rose sharply into adolescence. From birth, the chance of sensitization for any aeroallergen increased for each 12-month increment in age (OR ≥ 1.21, P < 0.01), with the largest effect observed for dust mite (OR = 1.56, P < 0.001). A steeper age-related rise in IgE antibody titre to dust mite, but no other allergen was associated with more severe disease. Despite this, sensitization to cat was more strongly associated with wheeze (OR = 4.5, P < 0.01), and linked to Fel d 1 and Fel d 4, but not Fel d 2. Comparison of cat allergic children with AD to those without, revealed higher IgE levels to Fel d 2 and Fel d 4 (P < 0.05), but not Fel d 1. CONCLUSIONS AND CLINICAL RELEVANCE: Differences in sensitization to cat and dust mite among young children with AD may aid in identifying those at increased risk for disease progression and development of asthma. Early sensitization to cat and risk for wheeze among children with AD may be linked to an increased risk for sensitization to a broader spectrum of allergen components from early life. Collectively, our findings argue for early intervention strategies designed to mitigate skin inflammation in children with AD.

The role of allergy in severe asthma.

The classification of asthma to identify forms which have different contributing causes is useful for all cases in which the disease requires regular treatment, but it is essential for the management of severe asthma. Many forms of the disease can occur, and complex mixtures are not uncommon; here we artificially separated the cases into four groups: (i) inhalant allergy, (ii) fungal sensitization with or without colonization (including ABPA); (iii) severe sinusitis with or without aspirin-exacerbated respiratory disease (AERD), and (iv) non-inflammatory cases, including those associated with severe obesity and vocal cord dysfunction (VCD). The reason for focusing on these groups is because they illustrate how much the specific management depends upon correct classification. Inhalant allergy can present as chronically severe asthma. However, severe attacks of asthma requiring hospital admission can occur in cases which are generally only mild or moderate. The best recognized and probably the most common cause of these acute episodes is acute infection with a rhinovirus. Recent evidence suggests that high titre IgE, particularly to dust mite, correlates to exacerbations of asthma related to rhinovirus infection. Although it is well recognized that the fungus Aspergillus can colonize the lungs and cause severe disease, it is less well recognized that those cases may not have full criteria for diagnosis of ABPA or may involve other fungi. Identifying fungal cases is important, because treatment with imidazole antifungals can provide significant benefit. Taken together, specific treatment using allergen avoidance, immunotherapy, anti-IgE, or antifungal treatment is an important part of the successful management of severe asthma, and each of these requires correctly identifying specific sensitization.

International consensus on (ICON) pediatric asthma.

Asthma is the most common chronic lower respiratory disease in childhood throughout the world. Several guidelines and/or consensus documents are available to support medical decisions on pediatric asthma. Although there is no doubt that the use of common systematic approaches for management can considerably improve outcomes, dissemination and implementation of these are still major challenges. Consequently, the International Collaboration in Asthma, Allergy and Immunology (iCAALL), recently formed by the EAACI, AAAAI, ACAAI, and WAO, has decided to propose an International Consensus on (ICON) Pediatric Asthma. The purpose of this document is to highlight the key messages that are common to many of the existing guidelines, while critically reviewing and commenting on any differences, thus providing a concise reference. The principles of pediatric asthma management are generally accepted. Overall, the treatment goal is disease control. To achieve this, patients and their parents should be educated to optimally manage the disease, in collaboration with healthcare professionals. Identification and avoidance of triggers is also of significant importance. Assessment and monitoring should be performed regularly to re-evaluate and fine-tune treatment. Pharmacotherapy is the cornerstone of treatment. The optimal use of medication can, in most cases, help patients control symptoms and reduce the risk for future morbidity. The management of exacerbations is a major consideration, independent of chronic treatment. There is a trend toward considering phenotype-specific treatment choices; however, this goal has not yet been achieved.

Risk factors for acute wheezing in infants and children: viruses, passive smoke, and IgE antibodies to inhalant allergens.

OBJECTIVE: To examine the prevalence of viral infection, passive smoke exposure, and IgE antibody to inhaled allergens in infants and children treated for acute wheezing. DESIGN: Case-control study of actively wheezing children who were compared with children without respiratory tract symptoms. SETTING: University of Virginia Pediatric Emergency Room. PATIENTS: Convenience sample of 99 wheezing patients (2 months to 16 years of age) and 57 control patients (6 months to 16 years of age). MEASUREMENTS AND RESULTS: Serum IgE antibody to inhalant allergens, measured by radioallergosorbent test (RAST), was uncommon in wheezing and control patients under age 2. After 2 years of age, the percentage of RAST-positive patients increased markedly and was significantly higher in wheezing patients than controls after age 4 (72%, n = 54, and 30%, n = 40, respectively, P < .001). Total IgE levels and nasal eosinophilia were strongly correlated with a positive RAST after age 2. Viral pathogens, predominantly respiratory syncytial virus, were identified in nasal washes from 70% (n = 20) of wheezing patients younger than 2 years of age compared with 20% of controls (n = 10), P < .05. After age 2, viruses, particularly rhinovirus, were isolated in washes from 31% (n = 70) of wheezing patients, 64% of whom were also RAST-positive. Levels of cotinine, a nicotine metabolite, were elevated (> or = 10 ng/mL) in saliva from a large percentage of smoke-exposed, wheezing patients under 2 (74%, n = 19) compared with those over 2 (14%, n = 51), P < .001. Odds ratios for wheezing were significant for virus (8.2, confidence interval [CI] = 1.3 to 5.0), and cotinine (4.7, CI = 1.0 to 21.3) in children under 2, and IgE antibody by RAST (4.5, CI = 2.0 to 10.2), virus (3.7, CI = 1.3 to 10.6), and the combination of IgE antibody and virus (10.8, CI = 1.9 to 59.0) were significant risk factors after age 2. CONCLUSION: Wheezing children younger than 2 years of age had a high rate of viral infection and a low rate of IgE antibody to inhalant allergens. When these children were exposed to passive smoke, salivary cotinine levels were elevated suggesting heavy exposure. After 2 years of age, sensitization to inhaled allergens became increasingly important and viruses remained a significant risk factor for wheezing. These data support recommendations to reduce tobacco smoke exposure at home, especially for young patients, and to consider sensitization to inhaled allergens and allergen avoidance in wheezing children at an early age, particularly after age 2 years.

Specific and nonspecific obstructive lung disease in childhood: causes of changes in the prevalence of asthma.

Reversible airway obstruction in childhood includes two major groups of patients: those with recurrent wheezing following bronchiolitis in early childhood, and those with allergic asthma, which represents an increasingly large proportion of cases through the school years. Over the last 40 years of the 20th century, allergic asthma has increased in many countries and in relation to several different allergens. Although this increase has differed in magnitude in different countries and also in the social groups most affected, it has had several features in common. The increase generally started between 1960 and 1970, has been progressive since then, and has continued into the 1990s without a defined peak. Among children 5-18 years of age, the increase has predominantly been among allergic individuals. Theories about the causes of the increase in asthma have focused on two scenarios: a) that changes in houses combined with increased time spent indoors have increased exposure to relevant allergens, or b) that changes in diet, antibiotic use, immunizations, and the pattern of infections in childhood have led to a change in immune responsiveness such that a larger section of the population makes T(H)2, rather than T(H)1 responses including IgE antibodies to inhalant allergens. There are, however, problems with each of these theories and, in particular, none of the proposed changes can explain the progressive nature of the increase over 40 years. The fact that the change in asthma has much in common with epidemic increase in diseases such as Type II diabetes or obesity suggests that similar factors could be involved. Several lines of evidence are reviewed that suggest that the decline in physical activity of children, particularly those living in poverty in the United States, could have contributed to the rise in asthma. The hypothesis would be that the progressive loss of a lung-specific protective effect against wheezing has allowed allergic children to develop symptomatic asthma. What is clear is that current theories do not provide either an adequate explanation of the increase or a practical approach to reversing the current trend.

Salivary cotinine levels in children presenting with wheezing to an emergency department.

We set out to evaluate salivary cotinine concentrations to judge tobacco smoke exposure among infants and children, and to examine the results in relation to age and wheezing. This was a case-control study of wheezing children (n = 165) and children without respiratory tract symptoms (n = 106) who were enrolled in the Pediatric Emergency Department at the University of Virginia. The age range of both wheezing and control patients was 2 months to 16 years. Questionnaires were combined with cotinine assays in saliva to evaluate exposure to environmental tobacco smoke (ETS) for each child. The prevalence of exposure to one or more smokers at home was high (68%); and 43% of the children enrolled were exposed to ETS from their mothers. According to the questionnaires, and after adjusting for age and race, a wheezing child in this study was more likely than a control to be exposed to at least one smoker at home (odds ratio = 1.9; 95% CI = 1.1-3.4). However, the odds of exposure to ETS from smoking mothers did not differ significantly between wheezing and control patients, and no significant association was found between the presence of wheezing and salivary cotinine levels. Among children exposed to ETS at home, cotinine levels were significantly higher in saliva from those under the age of two years, and from toddlers aged 2 and 3 years, compared to values from children over age 4 years. Moreover, the number of smokers in the home strongly influenced cotinine levels from children under age 4 years. In addition, higher cotinine levels were observed in saliva from children under age 2 years who were exposed to ETS from their mothers. Cotinine levels were similar and significantly correlated in paired samples of saliva and serum from children under 4 years of age (n = 54), (r = 0.92, P < 0.001). Based on information gathered from questionnaires, the results indicate that wheezing children were more likely than controls to be exposed to ETS at home. However, significant differences in ETS exposure between wheezing and control groups with respect to maternal smoke exposure or comparisons of salivary cotinine levels were not apparent. It was clear that determinations of salivary cotinine for monitoring the prevalence and intensity of household smoke exposure in this study were most valuable during the first 4 years of life.

Recent progress in mite allergen immunochemistry.

The successful purification of several mite allergens within the last few years has considerably enhanced our understanding of mite allergen immunochemistry. The role of these glycoproteins in stimulating human IgE ab and their role as immunogens in mice and rabbits has been studied extensively in a number of laboratories worldwide. In particular, purified allergens have facilitated the production of murine IgG Mabs that have been used to purify mite allergens by affinity chromatography; to investigate the diversity of antigenic sites on purified allergens; and to develop Mab based immunoassays for measuring allergen concentrations in dust samples and extracts. Full amino acid sequencing of several mite allergens is now in progress together with efforts to identify antigenically important peptide fragments. Such investigations are aimed to further increase our knowledge of humoral and cellular immune responses at the molecular level. For years, pollen counts have been used to judge airborne pollen allergen levels and to predict, in turn, the severity of symptom days for patients with hay fever. In contrast, simple methods for measuring dust allergens (e.g., mite allergen) have not been available. The development of Mab immunoassays, which can be converted from radiolabeled to enzyme labeled or fluorescence labeled assays, should provide rapid and quantitative measurements of specific mite allergen levels in house dust. Not only can such measurements provide useful clinical information in judging the exposure of patients to mite allergen, but the effectiveness of allergen avoidance regimes can be monitored objectively. By measuring the concentration of specific allergens in extracts, these assays could significantly improve efforts to standardize extracts used for diagnosis and treatment.

The relevance of allergen exposure to the development of asthma in childhood.

Sensitization to 1 or more of the common indoor allergens has been consistently associated with asthma among children and young adults (odds ratios for asthma, 3-18). For dust mite and cockroach allergens, there is a dose response relationship between domestic exposure and sensitization. Given that allergen provocation can induce many of the features of asthma, the findings strongly suggest that there is a causal relationship between allergen exposure in the home and asthma. However, it remains unclear at what time the critical exposure occurs (ie, in infancy or later) and what role allergen exposure has played in the increasing prevalence and severity of asthma. Objective evidence of an immune response to allergens is generally not present until after 2 years of age. Viral infections play several different roles in asthma in childhood. In infancy, respiratory syncytial virus infection can induce bronchiolitis and set up recurrent wheezing over the next few years. However, the risk factors for this are maternal smoking and small lungs at birth, rather than allergy. By contrast, the role of rhinovirus in precipitating attacks in children and young adults is strongly associated with allergy. Thus the likely scenario is that allergen exposure over the first few years of life induces sensitization (ie, T(H2) cells and IgE antibodies). Continuing exposure can maintain inflammation in the nose and lungs. However, many other factors contribute to wheezing such that there is no simple relationship between allergen exposure and asthma. Nonetheless, it is clear that the changes that have increased asthma have acted on allergic children.

Monitoring allergen exposure in asthma: new treatment strategies.

The development of monoclonal antibody-based enzyme-linked immunosorbent assay technology for measuring environmental allergen exposure has provided a benchmark for assessing the role of indoor allergens in causing asthma and other allergic diseases. Epidemiological studies from several parts of the world have shown that immunoglobulin E (IgE)-mediated sensitization to indoor allergens (mite, cat, dog and cockroach) is a risk factor for asthma attacks. A dose-response relationship between allergen exposure and sensitization has been demonstrated for mite allergens, and threshold values for exposure levels leading to sensitization or to exacerbations of symptoms have been defined. Comparative studies on airborne allergen levels have made it possible to determine the properties of aeroallergen particles, their concentration in indoor air, and the relationship to clinical symptoms. Together, these studies provide strong evidence that allergen exposure plays a causal role in the development of bronchial hyperreactivity and of the chronic inflammatory responses seen in patients with asthma. Logically, the primary preventive treatment should be allergen avoidance. Through knowledge of indoor allergen levels, both in dust and in the air, different avoidance strategies have been applied to the various indoor allergens, and there is increasing evidence of their clinical efficacy. Monitoring allergen levels in patients' houses should improve their understanding of the role of allergens in asthma and improve compliance with avoidance measures.

Airborne allergens associated with asthma: particle sizes carrying dust mite and rat allergens measured with a cascade impactor.

Patients with asthma may develop acute symptoms after exposure to domestic or laboratory animal allergens; however, they are usually not aware of a direct relationship between their acute attacks and exposure to pollen or dust mite allergens. The present experiments were designed to study whether the differences in symptoms could be explained by differences in the number or size of particles carrying airborne allergens. Airborne particles were collected with a filter or on the stages of a cascade impactor, and allergens were measured by use of inhibition radioimmunoassays. In rat rooms and during disturbance of rat litter, a large proportion of rat urinary allergen (45.9%) was collected on the second stage of the impactor (mean size approximately 7 microns diameter). When sampled 15 to 35 minutes after disturbance, 16% of these medium-sized particles were still airborne. By contrast, during disturbance of house dust, a significantly larger proportion of dust mite, antigen P1 (80.6 +/- 11.8%; p less than 0.001) was collected on the first stage of the impactor, and in keeping with the apparent size of these particles (diameter greater than 10 microns), very little of this allergen (less than 4%) was still airborne when sampled 15 to 35 minutes after disturbance. With nebulized diluted rat urine, approximately 75% of the allergen was collected on the fourth and final stages of the cascade impactor in keeping with the expected size, 0.5 to 3 microns in diameter. These results demonstrate that natural exposure to both allergens is strikingly different from the conditions used for bronchial provocation.(ABSTRACT TRUNCATED AT 250 WORDS)

Asthma and rhinitis related to laboratory rats: use of a purified rat urinary allergen to study exposure in laboratories and the human immune response.

Using the major rat allergen as a model has made it possible to study both natural exposure and the human immune response to an important laboratory animal. The close correlation between positive skin tests to whole rat urine and IgE antibody to the major urinary allergen in this and previous studies supports the use of this protein as a model rat allergen. Measurements of airborne rat allergen confirm that the maximum levels are higher than those reported with pollen or mite allergens. However, it is possible that exposure to rat allergens is comparable to levels of exposure to cat salivary allergens in houses with cats. The clear implication is that the high levels of exposure are responsible for the fact that a large proportion of exposed individuals develop IgG antibodies. Our results suggest that the prevalence of IgG antibodies (not individual levels) in a group of workers would be a good guide to exposure. This leaves unresolved why some of the individuals who develop IgG ab also develop IgE ab and become at risk for developing asthmatic responses. Only part of this risk is related to atopy. A striking feature of all the studies on animal allergy is the close association between IgE ab and asthma. It appears clear that it is those immune responses that include IgE ab that are a risk factor for asthma. It is not sensible for anyone to remain consistently sick with asthma and continue working with laboratory animals because there are well documented examples of occupational asthma that has not resolved after ceasing exposure.(ABSTRACT TRUNCATED AT 250 WORDS)

Epitope mapping of two major inhalant allergens, Der p I and Der f I, from mites of the genus Dermatophagoides.

The repertoire of antigenic sites on two major dust mite allergens, Der p I of Dermatophagoides pteronyssinus and Der f I of D. farinae, was studied using murine (BALB/c) monoclonal antibodies (Mab), polyclonal rabbit IgG antibodies, and human IgE antibodies. Fifty-three IgG Mab were analyzed from six different fusions (five vs Der p I, one vs Der f I). By antigen binding radioimmunoassay (RIA), most Mab were either Der p I or Der f I specific, and only 2/53 bound to both allergens. Epitope mapping studies using cold Mab to inhibit the binding of six 125I labeled Mab to solid phase allergen defined four nonrepeated, nonoverlapping epitopes on Der p I, a single species-specific epitope on Der f I and a cross-reacting epitope present on each allergen. All but one of the 53 Mab bound to one of these six epitopes. Seventy percent (25/35) of anti-Der p I Mab were directed to the same epitope, suggesting that this epitope is immunodominant for BALB/c mice. Similarly, 88% (16/18) of anti-Der f I Mab bound to the same epitope on Der f I. Parallel cross-inhibition curves were obtained using the species-specific Mab, 10B9, and the cross-reacting Mab, 4C1, to compete for binding to Der p I, suggesting that the epitopes defined by these two Mab on Der p I are adjacent to one another. Both murine Mab and polyclonal rabbit IgG antibodies to cross-reacting sites on both allergens were used to inhibit binding of human IgE antibodies to Der p I by using 19 sera from mite allergic patients. Cross-reacting rabbit IgG antibodies strongly inhibited all sera tested (mean 79.5% +/- 7.7) and two Mab, 10B9 and 4C1, partially inhibited (38% +/- 12). However, the four Mab directed against separate species-specific epitopes (including murine immunodominant sites) showed little or no inhibition (less than or equal to 20%). Our results suggest that most of the epitopes defined by Mab are not the same as, or close to, those defined by human IgE antibody. The striking differences in the repertoires of murine IgG and human IgE antibody responses to Der p I and Der f I could be explained by genetic differences or by altered antigen processing and presentation occurring as a result of different modes of immunization in mice and in mite allergic humans.

Changing concepts of allergic disease: the attempt to keep up with real changes in lifestyles.

In the past 100 years, changes have occurred in the outdoor environment and in houses that have contributed to the increased prevalence of hay fever and asthma. Much evidence indicates that exposure to indoor allergens is an important cause of asthma. Changes in housing that have contributed to the increased prevalence and severity of asthma include increased temperature, decreased ventilation, and permanent carpeting. In addition to these changes, geographic differences in allergens, deficiencies in cleanliness, poor health care, passive smoke, and lack of exercise also contribute to the increase in severity of asthma that has occurred. The management of asthma includes controlling exposure to indoor allergens and seeking additional treatable causes of asthma (e.g., fungal allergens). Changes will continue to occur, and physicians who treat allergic diseases should become involved in the design of houses to limit exposure. Many questions regarding allergen measurement and control remain.

Monoclonal antibodies to group II Dermatophagoides spp. allergens: murine immune response, epitope analysis, and development of a two-site ELISA.

BACKGROUND: Group II allergens are a major cause of sensitization in patients allergic to mites. To facilitate the antigenic analysis of group II allergens and to develop improved methods of allergen detection, we compared IgG anti-group II antibody responses in inbred mouse strains and raised a panel of monoclonal antibodies (mAbs). METHODS: IgE antibody responses were compared by antigen-binding radioimmunoassay. Epitope specificity of the mAbs was analyzed by two-site binding assays and by cross-inhibition radioimmunoassays. RESULTS: Comparison of polyclonal IgG antibody responses in five BALB congenic strains showed that H-2d mice had poor responses, whereas H-2b and H-2k mice had strong, cross-reactive, IgG anti-group II responses. The specificities of nine anti-Der p II IgE mAbs raised in A/J mice were compared with specificities of seven mAbs produced previously. Most mAbs (11 of 16) recognized common epitopes on Der p II and Der f II: three were specific to Der p II, and two showed high binding to Der f II. Epitope analysis showed that the mAbs defined four cross-reactive, nonoverlapping sites on the group II allergens. Binding of several combinations of mAbs was compared, and a two-site ELISA for group II antigens was developed. Linear regression analysis showed an excellent correlation between results of this assay and group II radioimmunoassay of house dust samples (n = 40, r = 0.85, p < 0.001). CONCLUSIONS: There are multiple cross-reactive B-cell epitopes on group II allergens. The group II ELISA has several important applications, including assessment of environmental allergen exposure, monitoring of the efficacy of avoidance procedures, and standardization of commercial mite allergen extracts.

The role of domestic allergens.

The documented increase in asthma has been almost entirely in perennial asthma and a large proportion of the cases are allergic to one of the common allergens found all year round in houses, i.e. house dust mites, cats, dogs or cockroaches. In population and case-control studies sensitization to one of these allergens is the strongest risk factor for asthma (adjusted odds ratios > or = 4). Using monoclonal antibody-based assays for the major indoor allergens it has been shown that sensitization to house dust mites is directly related to the concentration of Group 1 mite allergen in dust. This led to the hypothesis that increases in mite allergen secondary to changes in houses were responsible for increases in asthma. However, asthma has also increased in areas of the world where mites do not flourish. In these dry areas sensitization to one of the other indoor allergens is the major risk factor for asthma. Although sensitization of asthmatics reflects the concentration of allergens in their houses, these measurements of exposure do not accurately predict severity of symptoms. Other factors that can contribute to the symptoms of asthma may also have increased. In particular, diesel particulates, ozone, beta 2-agonists, endotoxin and rhinovirus infection have each been shown to enhance the inflammatory response to inhaled allergens. Increases in asthma must relate to some aspect of our predominantly sedentary indoor lifestyle; this could be either increased exposure to allergens or an increase in factors that enhance the response of the lungs to foreign proteins.

Airborne dust mite allergens: comparison of group II allergens with group I mite allergen and cat-allergen Fel d I.

The form in which allergens become airborne is important because it may influence both symptoms caused by allergen exposure and methods used to reduce exposure. The group I allergens from dust mites only become airborne during disturbance and fall rapidly, which is in keeping with their being carried on fecal pellets. Their mean size is approximately 20 microns in diameter. By contrast, the cat-allergen Fel d I is airborne on particles varying from greater than 10 to less than 2 microns in diameter, some of which remain airborne even without disturbance. A second group of mite allergens, molecular weight 14,000, are equally important and are associated predominantly with mite bodies. With a monoclonal antibody-based assay and a cascade impactor, we have investigated the form in which group II mite allergens become airborne. The results reveal that these allergens only become airborne during disturbance and that they fall within 15 minutes. However, the mean size of particles carrying group II allergens appears to be slightly smaller than the mean size of particles carrying group I allergens. In addition, the quantities of group II allergen becoming airborne during disturbance (mean, 26 ng/m3) could not be explained by the quantity found in fecal particles. Thus, group II mite allergens become airborne in a form quite distinct from cat allergens and very similar to group I mite allergens; however, it appears unlikely that fecal particles are the main form in which group II allergens become airborne.

Antigen Der f I from the dust mite Dermatophagoides farinae: structural comparison with Der p I from Dermatophagoides pteronyssinus and epitope specificity of murine IgG and human IgE antibodies.

The physicochemical and antigenic properties of an allergen purified from Dermatophagoides farinae, Der f I, were compared with Der p I from Dermatophagoides pteronyssinus. On SDS-PAGE, Der f I migrated as a single polypeptide chain with the same m.w. as Der p I (24,000). Two isoallergenic peaks of Der f I were identified on preparative isoelectric focusing (pI 5.7 to 6.3 and pI 6.6 to 6.95). Fractions from each peak were shown to have an identical amino acid composition (which was similar but not identical to Der p I) and the same N-terminal amino acid sequence. There was a good correlation between quantitative intradermal skin tests to both purified allergens and to D. farinae extract in mite-allergic patients, with positive results when using as little as 10(-5) micrograms/ml of Der f I. The majority of sera with detectable IgE antibody to D. farinae also had IgE antibody to Der f I both among children (29/42 = 69%) and adults (55/63 = 87%). By RAST, there was an excellent correlation between IgE antibody to Der f I and Der p I in sera from 42 mite-allergic children (n = 0.94, p less than 0.001). Polyclonal IgG antibodies from six mice immunized with Der f I showed preferential binding to that allergen, and most monoclonal antibodies (16 of 18) raised against Der f I did not bind Der p I. However, two monoclonal antibodies from this fusion showed cross-reactive binding to both allergens. Immunoabsorption experiments, using D. pteronyssinus and D. farinae extracts coupled to Sepharose, showed that a large proportion of murine antibodies (74% to Der p I and 60 to 93% to Der f I) could not be absorbed by the heterologous extract on the immunosorbent. In contrast, in sera from seven mite-allergic patients, most of the specific IgE and IgG antibody (i.e., greater than or equal to 82%) was removed by either immunosorbent. Thus, Der f I and Der p I represent a homologous pair of major allergens which possess both cross-reacting and species-specific epitopes. The antibody response in mice immunized with either allergen in complete Freund's adjuvant was largely directed against species-specific epitopes, whereas in allergic humans, IgE- and IgG-specific antibodies bound predominately to cross-reacting epitopes.