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Notes: Background: Many asthmatics in tropical and subtropical areas have positive skin prick tests to both Dermatophagoides spp. and to the mite Blomia tropicalis. This may be due to recognition by IgE of cross-reactive allergens between the different mite species or because of sensitization to species-specific allergens. A 14-kDa Blomia tropicalis allergen, Blo t 5, has been cloned and shows 40% sequence homology with Der p 5. The aim of this study was to investigate reactivity to B. tropicalis in patients known to be sensitized to D. pteronyssinus and to assess allergenic activity and cross-reactivity of recombinant (r) Group 5 allergens amongst these patients, who live in the UK and who are not exposed to B. tropicalis in their homes.Methods: Patients (n = 19) with asthma and/or rhinitis were selected based on clinical history and a positive skin prick test to D. pteronyssinus extract and were compared with non-allergic skin test negative controls (n = 10). IgE antibody responses to Blomia tropicalis, Dermatophagoides pteronyssinus, rDer p 5 and rBlo t 5 were compared by quantitative intradermal skin testing using serial 10-fold dilutions of each allergen. End point titre was the highest dilution giving an 8 × 8 mm wheal at 15 min. IgE antibodies to Blomia tropicalis, Dermatophagoides pteronyssinus, rDer p 5 and rBlo t 5 were measured using RAST, CAP and RIA, respectively.Results: All 19 patients had positive skin tests to D. pteronyssinus at concentrations of 0.001 to 1 AU/ml and 10 were skin test positive to rDer p 5 at concentrations of 10−4 to 5 µg/ml. Positive intradermal tests to Blomia tropicalis were seen in 12/19 patients at concentrations of 0.002 to 2 µg/ml. However none of the patients had positive skin tests to rBlo t 5. Non-allergic controls were all skin test negative at the highest concentration of each allergen tested. All subjects had quantifiable specific IgE to D. pteronyssinus, but only two had IgE to B. tropicalis. IgE to Der p 5 was found in six patients, but no patients had IgE to Blo t 5.Conclusions: This study of patients naturally exposed to D. pteronyssinus but not to Blomia tropicalis, provides evidence for IgE mediated cross-reactivity between allergens produced by both mite species. The results suggest that the Group 5 allergens of D. pteronyssinus and B. tropicalis are species-specific.

Notes: A group of 20 mite allergic asthmatic children aged 6-12 years old. living in Sao Paulo, Brazil, was studied regarding their degree of sensitization to house dust mites and exposure to mite allergens in their homes. In 18 out of 20 houses at least one dust sample was obtained which contained 〉 10 μg Der p I/g of dust. The highest levels of Dermatophagoides pteronyssinm allergens, Der p I and Group II, were measured in bedding samples (geometric mean 38.4 and 36.6 μg/g, respectively), followed by bedroom floor, TV room and kitchen. Mite allergen levels in Brazilian houses were as high as those reported to be associated with sensitization and acute attacks of asthma in other parts of the world. In keeping with previous reports that D. farinae is rarely found in Brazil, Der fl was undetectable or found in very low levels (〈0.5 μg/g). Levels of cat allergen Pel d I of 〉 8 μg/g of dust were obtained only in 2 houses only. Cockroach allergen Bia g I was detected in five out of 20 houses. Levels of IgE antibodies to D. pteronyssinus were 〉 200 RAST U/ml in 19 out of 20 children (geometric mean 1588 RAST U/ml). IgE antibodies to cat, cockroach, A. fumigatus, ragweed and rye grass pollens were undetectable or 〈80 RAST U/ml. IgE antibodies to the mite Blomia tropicalix were also measured, and levels 〉200 RAST U/ml were observed in 13 out of 20 sera. Immunoabsorption studies demonstrated that the bulk of the IgE- antibody to B. tropicafis (64%) was to species-specific allergens and that 36% were cross-reactive with D. pteronyssinus. The implication of our results is that management of children with asthma in São Paulo should include skin testing for allergy to both Dermatophagoides and B. tropicalis as well as recommendations about environmental control of house dust mite exposure.

Notes: Background Cat allergy is unique among allergy to mammals in that the major allergen Fel d 1 is a uteroglobin-like protein and not a lipocalin. The biochemical spectrum of the cat allergens is thus uncertain, particularly with regard to the role that a cat lipocalin protein may play in sensitization to cats in allergic individuals.Objective To analyse cDNA encoding a lipocalin allergen and the corresponding recombinant allergen at both the molecular and immunological levels.Methods A submandibular salivary gland cDNA expression library was constructed and screened for clones producing IgE-binding polypeptides. cDNA encoding a lipocalin allergen and its corresponding recombinant allergen were analysed.Results An IgE binding molecule with high sequence identity to the boar salivary lipocalin and the horse lipocalin Equ c 1 allergen was isolated and designated, Fel d 4. Serum from 62.96% of cat-allergic subjects examined had measurable IgE antibody to Fel d 4 but typically at low levels. Despite this in 47% of sera the anti-Fel d 4 IgE titres were higher than the anti-Fel d 1 titres. IgE binding to the lipocalin allergen could be blocked by an allergen extract from cow and to a lesser degree by extracts from horse and dog.Conclusion Fel d 4 is a lipocalin allergen produced by the cat, which binds IgE at relatively high frequency in cat-sensitive individuals. The allergen provides not only a means for investigating differences in the immune response to lipocalin allergens from that found for other mammalian species but also an important reagent for the diagnosis of cat allergy.

Notes: Background Cats are an important source of indoor allergens. However, only two cat allergens, Fel d 1 and albumin, have been cloned and sequenced. IgE antibodies to Fel d 1 and albumin do not fully account for IgE responses to cat and there is good immunochemical evidence that cats produce other allergens.Objective To identify and define the molecular structure of the other potential cat allergens.Methods A cat skin cDNA library was screened using pooled serum obtained from five asthmatic patients which contained high levels of IgE antibody to cat dander. Selected cDNA clones were screened by plaque immunoassay and one cDNA clone, encoding cystatin, was expressed in E. coli. The three dimensional structure of cat cystatin was modelled using the SWISS-MODEL computer program.Results Three positive cDNA clones (A, B and C) were identified, two of which were fully sequenced. Clones A and C encoded the same 98 amino acid residue sequence which showed 79% and 75% homology with bovine and human cystatin A, respectively. The cat cystatin sequence contained the conserved cysteine protease inhibitor signature and two of three lipocalin motifs. By plaque immunoassay, 60–90% of cat allergic sera had IgE ab to the expressed cystatin clones. The cysteine protease inhibitor motif was also partially conserved in dog allergen sequences, Can f 1 and Can f 2, which are lipocalins. The recombinant protein was expressed in E. coli as an 11-kDa protein, corresponding to the predicted MW of cat cystatin. The three-dimensional structure of cat cystatin was modelled on human cystatin structures.Conclusion A newly identified allergen, cystatin (Fel d 3), has been cloned from cat skin and is a member of the cysteine protease inhibitor family.

Notes: Background Sensitization and exposure to indoor allergens are the major risk factors for asthma. It is possible that significant exposure to domestic allergens occurs outside the home.Objectives To investigate the levels of Can f 1 and Bla g 2 in the dust from carpeted floors and upholstered seats in public buildings and public transport and the airborne concentrations of Der p 1, Fel d 1, Can f 1 and Bla g 2 in schools and offices.Methods Can f 1 and Bla g 2 were measured in the dust collected by vacuuming a I m2 area of carpet, as well as upholstered seats in five schools, six hotels, four cinemas, six pubs, three buses and two trains. Dust was also collected from the bedroom carpet, living room carpet, mattress and sofa in 20 homes with and 20 homes without a dog in the same area. Personal airborne sampling (2 L/min) was conducted for 8 h in offices (n= 16) and classrooms (n= 9). In addition, airborne samples in schools were collected using a high volume pump (60 L/min) for 1 h in three classrooms immediately after the children vacated the school. Can f 1, Bla g 2, Der p 1 and Fel d 1 were assayed using a two–site monoclonal antibody–based ELISA.Results Can f 1 was detected in all dust samples from public places, ranging from 0.2 to 52.5 μg/g, Significantly higher levels were found in upholstered scats (geometric mean – GM 9.4 μg/g) than in carpets (GM 1.5 μg/g; P 〈 0.001), and levels of Can f 1 〉 10 μg/g were found in 40% of upholstered seats in public places. Can f 1 was significantly higher in upholstered seats in public places than in sofas in homes without a dog (GM 1.8 μg/g; P 〈 0.001). Detectable levels of Bla g 2 were found in all of the schools (GM 2.4 U/g, range 0.8–4.4 U/g). Bla g 2 concentration greater than 2U/g (provisional threshold level representing risk of sensitization) was measured in 65% of the classrooms sampled. Der p 1 and Bla g 2 were below the detection limit in all airborne samples. However, airborne Fel d 1 and Can f 1 were detected in schools and offices, albeit in low concentrations.Conclusions Upholstered seats from public places constitute a reservoir for the accumulation of dog allergen, and a source of exposure to Can f 1 inside public buildings or on public transport. Exposure to cockroach allergens in schools may be important for cockroach sensitized asthmatic children.

Notes: Successful hyposensitization to Dermatophagoides pteronyssinus in perennial childhood asthma was associated with a slight mean rise of serum IgG antibody to the mite antigen in contrast to a placebo group in whom this antibody fell slightly. No relationship was detected between the effect on asthma and the magnitude of this change, Nor was there any consistent effect on IgE or IgA antibody.There was a wide range of total serum IgE and IgE, IgG and IgA antibody to D. pteronyssinus before treatment; this level did not predict the effect of treatment. Some patients lacked IgA antibody.IgE antibody to timothy grass pollen was raised in some but not others. These levels did not change systematically during the study and improvement occurred in those who had this antibody as well as IgE antibody to D. pteronyssinus.IgG and IgE antibodies to D. pteronyssinus were significantly correlated in the pre-treatment samples.Lymphocyte thymidine uptake was lower after 8 weeks of treatment than in the control group, not only after stimulus with D. pteronyssinus antigen, in vitro, but also with antigen from Candida albicans. This was not a serum effect.The late bronchial provocation response was lost only in those with serum IgA less than the log mean for age.

Notes: Background and Objective Chimeric mouse/human monoclonal IgGl and IgG4 antibodies were developed against the house dust mite allergen Der p 2. These chimeric IgG antibodies, hIgG1-Dp2 A and hIgG4-Dp2 A, have the same binding characteristics as the previously reported chimeric hIgE-Dp2 A and are composed of the heavy chain variable domains and light chains ot the original murine monoclonal antibody 2B12., whereas the heavy chain constant domains have been replaced by the human IgGl or IgG4 heavy chain. The expression level of hIgG1-Dp2 A and hIgG4-Dp2 A was 1 and 3.5 μg/mL, respectively.Methods and Results Since all IgG in these culture supernatants is allergen-specific. they are useful reference reagents and enable the calculation of the amount of allergen specific IgG l and IgG4 antibodies in absolute IgG amounts. The results obtained with two panels of sera from patients in immunotherapeutic treatment were evaluated and compared in Der p 2 IgE, IgGl and IgG4 RAST and with reversed lgG4 RAST using labelled purified Der p 2. Close agreement between the results for the two IgG4 assays was found.Conclusion With these chimeric reference reagents the quantities of isotype specific antiallergen antibodies can be calculated and compared.

Notes: Background Sensitization and exposure to mile allergens is a major risk factor for asthma. Little is known about the rate of build-up of allergens in the mite microhabitats.Objectives To investigate the rate of increase in mite allergen levels in new mattresses.Methods Der p 1 was measured in the dust samples collected from six identical new single mattresses over a period of 2 years.Results Der p 1 increased significantly at 4 months as compared with baseline level (P 〈 0.01), but no difference was found between the concentrations at 4. 8, 12 and 24 months. There was a significant correlation between Der p 1 concentration in mattresses at 4.8, 12 and 24 months and Der p 1 levels in the bedroom carpet at the beginning of the study.Conclusions New mattresses can become a significant source of exposure to mite allergens after a short period of time (〈 4 months). There is little justification for advising mite sensitive patients to replace their mattresses as a part of avoidance regime.