A sensitive reverse ELISA for the measurement of specific IgE to Der p 2, a major Dermatophagoides pteronyssinus allergen.

BACKGROUND: Epidemiologic studies have shown that the presence of IgE antibodies to house dust mite and other indoor allergens is an important risk factor for asthma. OBJECTIVE: The aim of this study was to develop a reverse ELISA (rELISA) for measuring specific IgE to Der p 2, a major Dermatophagoides pteronyssinus (Dpt) allergen, as a potential tool for followup of allergen immunotherapy. METHODS: Recombinant Der p 2 allergen or a monoclonal antibody to Der p 2 was used to coat plates in conventional ELISA (cELISA) and rELISA, respectively. Sera from 48 asthmatic patients with positive skin prick test (SPT+) to D. pteronyssinus extract were analyzed for total IgE and specific IgE to Der p 2, and the results were compared with a group of 41 SPT asthmatic and 30 SPT- control subjects. RESULTS: The sensitivity of the two assays for Der p 2-specific IgE was 3.9 EU/mL and their specificities were confirmed by inhibition tests, in a dose-dependent manner. There was a significant positive correlation between cELISA and rELISA (r = 0.74; P < 0.0001). However, rELISA was more sensitive than was cELISA, regarding both the positive sera percentage (70.8% vs 52.1%) and the Der p 2-specific IgE levels (28.4 vs 4.5 EU/mL) in SPT+ asthmatic patients. CONCLUSIONS: rELISA has shown to be a sensitive and alternative method for measuring Der p 2-specific IgE without using radioactive techniques. Detection of specific IgE to major allergens and relevant peptides, and identification of B cell epitopes in allergens will provide valuable information for the design of allergen analogs and peptides for immunotherapy.     

Sensitization to Blomia tropicalis and dermatophagoides pteronyssinus-a comparative study between Singapore and Taiwan

Blomia tropicalis (Bt) and Dermatophagoides pteronyssinus (Dp) are the predominant domestic mites species in Singapore and Taiwan. This study aims to characterize and compare the mite sensitization profiles in both countries. Skin prick tests were performed on 203 Singaporeans with Dp and Bt crude extracts. In vitro IgE and IgG4 reactivity to extracts and specific allergens (Der p 1, Der p 2 Der p 5 and Blo t 5) were determined by immunoassays. Approximately 91% of the tested Singaporeans were skin test positive for both Bt and Dp. Both populations share similar frequencies of in vitro IgE reactivity to all the allergens tested, but they differ in the pattern and magnitude of allergen sensitization. Although Der p 1, Der p 2 and Blo t5 are major sensitizing allergens in both countries, Blo t 5 is a more potent one in Singapore, probably reflecting the high level of exposure to Bt. The unique major Bt and Dp allergens should be included for precise diagnosis and effective immuno-therapeutic treatment of mite allergy in both countries.     

Molecular cloning and characterization of full-length cDNAs encoding a novel high-molecular-weight Dermatophagoides pteronyssinus mite allergen, Der p 11

BACKGROUND: Dermatophagoides pteronyssinus (Dp) and D. farinae (Df) mites are the most important source of indoor aeroallergens. Most Dp mite allergens identified to date have relatively low molecular weights (MWs). Identification of high-MW mite allergens is a crucial step in characterizing the complete spectrum of mite allergens and to provide appropriate tools for diagnostic and therapeutic application. METHODS: The full-length Der p 11 cDNA clone was isolated using cDNA library immunoscreening, the 5'-3' rapid amplification of cDNA ends (RACE) system and polymerase chain reactions (PCR). The whole cDNA insert and its PCR-derived DNA fragments (p1 to p4) were generated and expressed in the Escherichia coli expression system. The allergenicity of the recombinant protein and its peptide fragments was examined by IgE immunodot assays. The IgE-binding reactivity of rDer p 11 was analyzed in the serum of 50 asthmatic children with positive reactivity to Dp mite extract. Its recombinant peptide fragments were also examined by immunodot assays in 30 mite-allergic children. RESULTS: Der p 11 cDNA consists of a 2625-bp open reading frame encoding a 103-kDa protein with 875 amino acids. It exhibits significant homology with the paramyosin of other invertebrates. The protein sequence alignment of this newly identified Dp mite allergen (denominated as Der p 11) revealed over 89% identity with Der f 11 and Blo t 1. Among 50 Dp-sensitive asthmatic children, rDer p 11 showed positive IgE-binding reactivity to 39 patients (78%). Using immunodot assays, multiple human IgE-binding activities were demonstrated in all four fragments of Der p 11. Using immunoblot assays, the dominant IgG-binding epitope for monoclonal antibody (mAb642) was located in fragment p3 only. In immunoblot assays, cross-inhibition between rDer p 11 and rDer f 11 was up to 73-80% at concentrations of 100 microg/ml. CONCLUSIONS: This study confirms that the newly identified recombinant Der p 11 is a novel and important high-MW Dp mite allergen for asthmatic children. Our data also indicates that human IgE-binding major epitopes are scattered over the entire molecule of Der p 11.     

An extensive study of human IgE cross-reactivity of Blo t 5 and Der p 5

BACKGROUND: Dual sensitization by Blomia tropicalis and Dermatophagoides pteronyssinus mites is common in tropical and subtropical countries. The human IgE cross-reactivity between clinical important group 5 allergens, Blo t 5 and Der p 5, remains controversial. OBJECTIVE: This study was undertaken to assess the levels of the IgE cross-reactivity between Blo t 5 and Der p 5 by using sera from a large cohort of asthmatic children in subtropical and tropical countries. METHODS: Purified recombinant Blo t 5 and Der p 5 were produced in Pichia pastoris and tested against sera from 195 asthmatic children. The IgE cross-reactivity was examined by direct, inhibitory and competitive human IgE enzyme-linked immunosorbent assay as well as skin prick tests. RESULTS: The Blo t 5 IgE responses were 91.8% (134 of 146) and 73.5% (36 of 49) for Taiwanese and Malaysian sera, respectively. The Blo t 5 specific IgE titers were significantly higher than those of Der p 5 (P <.02). The correlation of IgE reactivity between Blo t 5 and Der p 5 was low, and only limited cross-reactivity was observed. This was further confirmed by the dose-response inhibition studies. Skin prick tests performed on asthmatic children in Thailand also showed differential IgE response to Blo t 5 and Der p 5. CONCLUSION: By using a large panel of asthmatic sera and a combination of in vitro and in vivo assays, the major allergen of B tropicalis in tropical and subtropical regions, Blo t 5, exhibits low levels of IgE cross-reactivity with homologous Der p 5. These findings suggest that highly specific clinical reagents are necessary for precise diagnosis and immunotherapeutic treatment of sensitization to group 5 mite allergens.     

Sensitization to Dermatophagoides pteronyssinus and Blomia tropicalis extracts and recombinant mite allergens in atopic Thai patients

Mite surveys in Thailand indicated that Dermatophagoides pteronyssinus (Dp) is predominant, but so far there were no data available on Blomia tropicalis (Bt), which is prevalent in the Asia Pacific region. Skin prick testing (SPT) was performed in 40 atopic children, 45 atopic adults and 17 non-atopic volunteers. Skin reactions to Dp were found in 25/40 (62.5%) and 23/45 (51.1%); skin reactions to Bt were found in 15/40 (37.5%) and 18/45 (40%) in atopic children and adults, respectively. SPT to the major sensitizing allergens Der p 1, Der p 2, Der p 5, and Blo t 5 showed positive results in 14/40 (35%), 12/40 (30%), 1/40 (2.5%) and 4/40 (10%) of atopic children, and in 12/45 (26.7%), 13/45 (28.9%), 5/45 (11.1%), 6/45 (13.3%) of atopic adults, respectively. The results indicate that Dp is one of the major sources of allergy, while Bt is a minor one and that Der p 1 and Der p 2 are important mite allergens in Chiang Mai, Thailand.     

广州地区支气管哮喘患者常见吸入变应原调查分析

目的　调查广州地区支气管哮喘发病相关的主要致敏变应原 .方法　对 10 2例支气管哮喘缓解期患者 ,其中儿童 4 8例 ,成人 5 4例 ,用尘螨、霉菌、猫毛等 12种常见吸入变应原进行皮肤点刺试验 ,并测定其中 2 7例总IgE(TIgE)的水平 .结果　儿童组尘螨皮试阳性率最高 (均为 79.2 % ) ,其次是屋尘 (72 .9% ) .成年组尘螨皮试阳性率最高 ,粉尘螨和屋尘螨分别为 5 9.3%、6 2 .9% ,其次为屋尘 (4 0 .7% ) .在动物毛发皮屑中 ,以狗毛为主 ,猫毛次之 ;对于所有变应原 ,儿童组的阳性百分率远远高于成年组 ;2 7例血清总IgE的均值为 5 0 6 .1KU/L ,远高于正常人 ,其中皮试阳性者TIgE水平高于皮试阴性者 (p<0 .0 1) .结论　广州地区主要变应原均为尘螨 ,且哮喘儿童对尘螨过敏的阳性率显著高于成人     

Use of a chimeric ELISA to investigate immunoglobulin E antibody responses to Der p 1 and Der p 2 in mite-allergic patients with asthma, wheezing and/or rhinitis

BACKGROUND: Sensitization to indoor allergens, particularly to dust mites, is a strong risk factor for asthma in children and adults. Assessment of sensitization is carried out using in vivo and in vitro tests to detect specific IgE antibodies. OBJECTIVE: To investigate IgE antibody responses to mites in patients with asthma, wheezing and/or rhinitis, using chimeric ELISA to measure specific IgE antibodies to mite allergens Der p 1 and Der p 2. METHODS: Specific IgE antibodies to Der p 1 and Der p 2 were quantified by chimeric ELISA, and compared with IgE to Dermatophagoides pteronyssinus (Dpt) measured using the CAP system (Pharmacia). A panel of sera from 212 patients with asthma, wheezing and/or rhinitis and 11 controls was analysed. RESULTS: There was a significant correlation between IgE to Dpt measured by CAP and IgE to Der p 1 (r = 0.81, P < 0.001), Der p 2 (r = 0.79, P < 0.001) and combined Der p 1 and Der p 2 (r = 0.86, P < 0.001). Seventy per cent of all patients had IgE to Dpt, and of those, 76.5% had IgE to Der p 1, 79.2% had IgE to Der p 2 and 83.1% had IgE to Der p 1 and Der p 2 combined. Considering the cut-off level of 2 IU/mL of IgE to either Der p 1 or Der p 2, the predictive value for a positive IgE to Dpt by CAP was greater than 95%. CONCLUSIONS: The chimeric ELISA allowed accurate quantification of IgE antibodies to Dpt allergens Der p 1 and Der p 2, and it could be useful for studying immune responses to mites in patients with asthma and/or rhinitis.     

Buckwheat anaphylaxis: an unusual allergen in Taiwan

IgE-mediated hypersensitivity to buckwheat is common in Korea, Japan, and some other Asian countries. However, buckwheat is not a common allergen in Taiwan. We report a woman with asthma who had anaphylactic shock, generalized urticaria, and an acute exacerbation of asthma five minutes after ingesting buckwheat. The patient underwent skin prick and Pharmacia CAP testing (Uppsala, Sweden) for specific IgE to buckwheat, white sesame and soybean as well as other common allergens in Taiwan including Dermatophagoides pteronyssinus (Dp), D. farinae (Df), cat and dog dander, cockroach, egg white, cow milk and codfish. The patient had a strongly positive skin prick test response to buckwheat and positive reactions to Dp and latex. Specific IgE results were class 6 for buckwheat, class 4 for Dp and Df, and class 2 for dog dander, wheat, sesame and soybean. Results of an open food challenge with white sesame and soybean were negative. Although buckwheat is a rare allergen in Taiwan, it can cause extremely serious reactions and should be considered in patients presenting with anaphylaxis after exposure to buckwheat.     

Immune reactivity to Der p I and Der p II in house dust mite sensitive patients attending paediatric and adult allergy clinics

A comparison was made of immune responses to house dust mite allergens in symptomatic patients from paediatric and adult allergy clinics. IgE-specific immune reactivity to Dermatophagoides pteronyssinus was examined by Western blotting and, for Der p I and Der p II. by radioimmune dot-blot using purified allergens. Nineteen mite-sensitive children (mean age 9 years) and 26 adults (mean age 31 years) were compared. Positive IgE responses by dot-blot were found to Der p I and Der p II in 79% of children, whereas reactivity was only present in 23% and 19% respectively of adults, and densitometry indicated a weaker response. In children. Western blotting indicated that the majority of the serologic reactivity was directed to Der p II (17/19, 89%) or a 100 kDa fraction whereas in adults, reactivities were generally directed to other fractions with only 15/26 (58%) recognizing Der p II. Consistent with results of others, a fraction corresponding to Der p I was poorly detected on Western analysis despite positive dot-blots. To determine whether the relative lack of IgE serological immune reactivity in adults was associated with a similar lack of cellular recognition, T-cell proliferation studies were performed, also using purified allergens. Interestingly, these revealed that cellular responsiveness, without serological reactivity, was present in 29% of subjects to Der p I and 50% to Der p II. Proliferative responses were evident in all individuals with specific IgE.     

HLA DPB1*0201 allele is negatively associated with immunoglobulin E responsiveness specific for house dust mite allergens in Taiwan

BACKGROUND: House dust mite (HDM) Dermatophagoides pteronyssinus is the most important source of indoor allergens that cause allergic diseases in Taiwan. We prepared purified HDM allergens (Der p 1, Der p 2 and Der p 5) to detect allergen-specific immunoglobulin (Ig) E responsiveness among a large number of test subjects. The robust genetic typing system for HLA class II genes also facilitated the study on association of HLA and allergic response toward HDM. OBJECTIVE: This study intended to investigate the association between HLA class II alleles and the IgE responsiveness to the major allergens from HDM, D. pteronyssinus. METHODS: Two hundred and forty-eight subjects were selected for HLA association study. Plasma HDM allergen (Der p 1, Der p 2, Der p 5) -specific IgE and Der p 2-specific IgG antibodies were detected by ELISA, while HLA class II -DRB1, -DQA1, -DQB1, -DPB1 genetic polymorphism was determined by polymerase chain reaction/sequence-specific oligonucleotide probe hybridization (PCR/SSOPH). Statistical comparison of the allelic distribution of each HLA class II genes among the individuals with/without HDM allergen-specific IgE and IgG antibodies were performed. RESULTS: There was no significant association between HLA DRB1, DQB1, DQA1 alleles and HDM-specific IgE responsiveness noted. Only DRB1*0803 and the linked DQA1*0103 alleles showed positive association with Der p 5-specific IgE responsiveness. However, we found that HLA-DPB1*1301 predisposed subjects to IgE responsiveness to HDM Der p 5. HLA DPB1*0501 was weakly associated with the IgE responsiveness to HDM Der p 1 and Der p 5. There was a strong negative association between the HLA-DPB1*0201 allele with IgE responsiveness to Der p 1 (OR: 0.30, P 相似文献   

Higher incidence of Dermatophagoides pteronyssinus allergy in children of Taipei city than in children of rural areas.

BACKGROUND AND PURPOSE: House dust mites are the most common cause of sensitization in wheezing children in most parts of the world. The aim of this study was to investigate prevalence of sensitization and the levels of specific immunoglobulin E (IgE) to Dermatophagoides pteronyssinus (Der p) in wheezing children in Taipei city and central rural Taiwan. METHODS: A total of 3546 children were enrolled in this study. Children were grouped into those living in Taipei city (n = 1340) and those residing in the rural part of central Taiwan (2206). The prevalence of sensitization and level of specific IgE antibody to Der p and cockroach were analyzed by age and geographic area. RESULTS: The results showed significantly higher sensitization rates and mean specific IgE levels to Der p with increasing age of patients in both Taipei city and rural central Taiwan. In addition, children from Taipei city had a significantly higher average sensitization rate of Der p than rural children (p<0.05). The proportion of children sensitized to cockroach increased with age both in Taipei city and central rural Taiwan, but the specific IgE levels to cockroach were not statistically different between the 2 groups (p=0.061). CONCLUSIONS: Sensitivity to aeroallergens varies with age and with geographical location (city vs rural) in asthmatic children. Circulating IgE antibodies against Der p were common at all ages. Cockroach allergen is also associated with recurrent wheezing in Taiwan. Avoidance of indoor aeroallergens such as Der p and cockroach allergen should be an important component in plans for the management of recurrent wheezing.     

Allergenic differences between the domestic mites Blomia tropicalis and Dermatophagoides pteronyssinus

BACKGROUND: House dust mite allergens are the most important indoor allergens associated with asthma and rhinitis in Singapore and the tropics. Recent data to suggest that besides the Dermatophagoides spp., the domestic mite Blomia tropicalis (Bt) is also an important source of allergens in these regions. OBJECTIVE: To evaluate the degree of allergenic cross-reactivity between Bt and D. pteronyssinus (Dp). METHODS: Cross-reactivity between extracts of Bt and Dp was evaluated by fluorescent allergosorbent (FAST) inhibition studies and cross enzyme immunoelectrophoresis. Additionally, the major Dp allergens - Der p 1, Der p 2 and Der p 5, were also compared with the Bt extract by dot blot inhibition. Skin prick and intradermal end-point titration were then carried out to compare the homologous allergens of the mite species, Blo t 5 and Der p 5. RESULTS: FAST inhibition studies showed low to moderate cross-reactivity between the two dust mite extracts (maximum cross-inhibition, 60%). Native allergens studied by cross enzyme immunoelectrophoresis using mite allergic sera also showed similar results but with at least four cross-reactive IgE binding antigens. Dot blot inhibition studies using allergens of Dp, Der p 1, Der p 2, and Der p 5, showed little cross-reactivity between these allergens with components of the crude Bt extracts. Further, evaluation of a recombinant allergen of Bt, Blo t 5, showed low levels of cross-reactivity even with its homologous Dp counterpart, Der p 5. CONCLUSION: These results provide evidence that Bt allergens are distinct and have relatively low to moderate cross-reactivity with Dermatophagoides spp. allergens. Bt allergens should therefore be included in the diagnostic panel for the evaluation of allergic disorders in the tropics, and the development of new diagnostic and therapeutic strategies should include allergens of Bt.     

Protein sequence analysis of a novel 103-kDa Dermatophagoides pteronyssinus mite allergen and prevalence of serum immunoglobulin E reactivity to rDer p 11 in allergic adult patients

BACKGROUND: House dust mites are regarded as important indoor allergens. While the most studies mite allergens are low molecular weight (mw), a high mw Dermatophagoides farinae mite paramyosin (Der f 11) has recently been cloned. We have also cloned a novel high mw Dermatophagoides pteronyssinus (Dp) mite allergen, Der p 11. OBJECTIVE: The aim of this study was to isolate and express a cDNA gene coding for a Der p 11 allergen, to compare the sequence of Der p 11 with other antigens and to evaluate the presence of IgE reactivity to the recombinant protein (rDer p 11) in the sera of allergic adult patients. METHODS: The full-length Der p 11 gene was isolated by cDNA library screening, 5'-3' rapid amplification of cDNA ends and PCR. The cDNA gene was expressed as a glutathione-S-transferase fusion protein in Escherichia coli. The allergenicity of rDer p 11 was tested by human IgE immunodot or immunoblot assay in a large panel of 100 allergic patients with bronchial asthma, allergic rhinitis or eczema. RESULTS: Der p 11 is a 2965 bp cDNA gene with a 2625 bp open reading frame coding for a 875 amino acid protein. The deduced amino acid sequence of the Der p 11 showed significant homology with various invertebrate paramyosins. The prevalence of serum IgE reactivity to rDer p 11 on immunodot assay ranged from 41.7% to 66.7% in different allergic patient groups, whereas it was rare in non-atopic patients with urticaria (18.8%) and in normal individuals (8%). A high frequency (five out of eight) of MAST(Dp)- allergic serum samples had specific IgE-binding activity to rDer p 11 or its fragments on immunoblot assay, even though their IgE-binding activity to Dp extract was either weak or negative. CONCLUSION: The 103-kDa Der p 11 appears to be major Dp mite allergen with a high frequency of IgE reactivity in sera of patients allergic to mites.     

Quantitative measurement of IgE antibodies to purified allergens using streptavidin linked to a high-capacity solid phase

BACKGROUND: Commercially available assays for IgE antibody provide results in international units per milliliter for many allergen extracts, but this is not easily achieved with purified or novel allergens. OBJECTIVE: To develop assays for IgE antibody suitable for purified or novel allergens by using a commercially available immunosorbent. METHODS: Streptavidin coupled to a high-capacity immunosorbent (CAP) was used to bind biotinylated purified allergens from mite (Der p 1 and Der p 2), cat (Fel d 1), and dog (Can f 1). Assays for IgE antibody to these allergens were performed on sera from children (asthma and control) as well as adults with atopic dermatitis. RESULTS: The results were validated by serial dilution of sera with high and low levels of IgE antibody and were quantitated in international units per milliliter by using a standard curve. Values for IgE antibody to Der p 1, Der p 2, and Fel d 1 correlated with values obtained with the allergen extracts (r2 = 0.80, 0.84, and 0.95, respectively; P < .001 in each case). Furthermore, the values for IgE antibody in sera from children with high exposure to mite and cat allergens demonstrated 10-fold higher levels of IgE antibody to Der p 1 and Der p 2 than to Fel d 1 (P < .001). CONCLUSION: The streptavidin immunosorbent technique provides a new method for quantifying IgE antibody to purified proteins. The results provide evidence about the high quantities of IgE antibody to purified inhalant allergens in patients with atopic dermatitis. In addition, the results demonstrate major differences in IgE antibodies specific for mite and cat allergens among children with high exposure to both allergens.     

Sensitization profiles of Malaysian and Singaporean subjects to allergens from Dermatophagoides pteronyssinus and Blomia tropicalis

BACKGROUND: The house dust mites Dermatophagoides pteronyssinus (Der p) and Blomia tropicalis (Blo t) are the most common house dust mite species in Southeast Asia. To date, there have only been a few studies on the sensitization profile of the general populations in Southeast Asia to house dust mites. The aim of this study was to determine the profiles of Der p and Blo t sensitization among Singaporean and Malaysian subjects. METHODS: Enzyme-linked immunosorbent assay was used to detect specific IgE to Der p and Blo t mite crude extracts as well as purified Der p 1, Der p 2 and Blo t 5 allergens. Sera used were from 229 Singaporean subjects (124 with rhinitis, 105 without rhinitis) and 143 Malaysian subjects (94 adults and 49 children with asthma). RESULTS: The sensitization profile of rhinitis subjects to the dust mite allergens used in this study was as follows: Blo t extract positive: 91/124 (73%); Blo t 5 positive: 62/124 (50%); Der p extract positive: 61/124 (49%); Der p 1 positive: 53/124 (43%); Der p 2 positive: 45/124 (36%). The nonrhinitis subjects' sensitization profile was as follows: Blo t extract positive: 60/105 (57%); Blo t 5 positive: 24/105 (23%); Der p extract positive: 38/105 (36%); Der p 1 positive: 14/105 (13%); Der p 2 positive: 17/105 (16%). The study of Malaysian asthmatic adults showed that 39% of them were sensitized to Der p 1, 32% to Der p 2 and 37% to Blo t 5. Among the asthmatic children, sensitization to Blo t 5, Der p 1 and Der p 2 was 90, 57 and 39%, respectively. CONCLUSION: This study clearly revealed that dual sensitization to B. tropicalis and D. pteronyssinus is common in the general populations of Singapore and Malaysia. Sensitization to Blo t 5 is more prevalent than to Der p 1 and Der p 2.     

Indoor mite allergens in patients with respiratory allergy living in Porto, Portugal

We investigated the levels of mite allergens (Der p 1, Der f 1, Der 2, and Lep d 1) in dust samples from the homes of 59 patients with asthma, 36 sensitized to house-dust mites (HDM) and 23 to grass pollen (controls), living in Porto, northern Portugal. The relationship between exposure and sensitization to HDM and the influence of housing conditions on mite-allergen levels were also evaluated. Der p 1 (median 9.2 μg/g) and Der 2 (4.6 μg/g) were the main allergens, while Der f 1 and Lep d 1 levels were always <1 μg/g dust and undetectable in 11% and 47% of samples, respectively. All HDM-sensitized asthmatics were exposed to Der p 1 levels >2 μg/g and their homes contained significantly higher levels of Der p 1 (median 12.5 vs 6.4 μg/g; P=0.008) and Der 2 (6.2 vs 3.0 μg/g; P=0.004) when compared to the control group. A significant correlation was observed between the exposure to Der p 1 and the wheal area at skin testing with the Dermatophagoides pteronyssinus (Dp) extract (P=0.01) as well as with serum specific IgE levels to Dp (P=0.03). Patients with higher levels of serum specific IgE (≥17.5 HRU/ml) were also more frequently exposed to Der p 1 levels ≥10 (μg/g (P=0.002). Old homes, presence of carpets, and signs of dampness were conditions associated with significantly higher levels of mite allergens. In conclusion, we found high levels of Der p 1 and Der 2 particularly in the homes of HDM-sensitized patients and we confirm the relationship between exposure and sensitization to HDM, assessed by both in vivo and in vitro methods. In addition to a favorable outdoor climate, we found in our region housing conditions propitious to mite growth, suggesting that specific geographic characteristics must also be taken into account for the correct planning of mite-avoidance measures.     

The chitinase allergens Der p 15 and Der p 18 from Dermatophagoides pteronyssinus

BACKGROUND: House dust mites Dermatophagoides pteronyssinus and Dermatophagoides farinae cause allergic disease in humans as well as in dogs. In geographical regions where the two mite species coexist, they both elicit specific immunoglobulin (Ig E) responses in humans whereas dogs preferentially react to D. farinae extracts. In dogs the main IgE binding is directed to the D. farinae chitinase allergens Der f 15 and Der f 18 and not to the groups 1 and 2 allergens as found for humans. Although the IgE response of humans to Der f 18 has been investigated there is no report on Der f 15-specific IgE in humans. OBJECTIVE: This study aimed to characterize the chitinase allergens Der p 15 and Der p 18 of D. pteronyssinus and to find out whether they are important allergens for humans. METHODS: cDNA was cloned by a polymerase chain reaction strategy from D. pteronyssinus libraries using primers based on conserved chitinase sequences. IgE binding to the recombinant polypeptides was measured by immunosorbent assay. Mice were immunized with the polypeptides and cross-reactivity examined. RESULTS: Two variants of Der p 15 were isolated, encoding mature proteins of 58.8 and 61.4 kDa. The amino acid sequences had 90% identity to Der f 15. The cDNA for Der p 18 encoded a mature protein of 49.2 kDa with 88% sequence identity to Der f 18. Der p 15-specific IgE was detected in 70% and Der p 18-specific IgE in 63% of a panel of 27 human allergic sera. CONCLUSIONS: The D. pteronyssinus chitinases Der p 15 and Der p 18 show a high frequency of binding to IgE in allergic human sera. They are therefore potentially important allergens for humans as well as dogs.     

Characterization of glutathione S-transferase from dust mite, Der p 8 and its immunoglobulin E cross-reactivity with cockroach glutathione S-transferase

BACKGROUND: Sensitization to mite and cockroach allergens is common, and diagnosis and therapy of allergy can be further complicated by the presence of allergen isoforms and panallergens. Purified recombinant and native allergens are useful for studies to resolve such problems. OBJECTIVE: To assess the allergenicity of native and recombinant mite glutathione S-transferase (GST) (Der p 8) and study the IgE cross-reactivity between Der p 8 and cockroach GST. METHODS: Der p 8 cDNA encoding a new isoform was isolated and expressed in yeast. Native Der p 8 was affinity purified from mite extract. IgE reactivity to native and recombinant Der p 8 was assessed by ELISA using sera from allergic subjects from Taiwan, Singapore and Malaysia. IgE cross-reactivity between Der p 8 and cockroach GST was examined by IgE inhibition assays. RESULTS: Our Der p 8 cDNA encoded a basic isoform (pI=8.5) containing six polymorphic residues located at positions 46, 106, 149, 160, 167 and 184. At least 8 isoforms of native Der p 8 were detected by two-dimensionalgel and immunoblot analyses. Sera from Taiwanese asthmatics showed 96% and 84% IgE reactivity to native Der p 8 and recombinant Der p 8, respectively. Native Der p 8 showed 75% and 65% IgE reactivity with sera from Malaysia and Singapore, respectively. CONCLUSIONS: A high frequency of sensitization to mite GST among allergic subjects was observed but the titres of IgE reactivity were low. The IgE cross-reactivity between mite and cockroach GST suggests that GST is a panallergen.     

IgE responses to Dermatophagoides pteronyssinus native major allergens Der p 1 and Der p 2 during long-term specific immunotherapy

We investigated by ELISA the IgE response to whole extract of the house-dust mite Dermatophagoides pteronyssinus (Dp) and to the native major allergens, Der p 1 and Der p 2, in sera from 18 adult patients (group A) with Dp-allergic asthma before ( t ₀) and 1, 2, 3, and 4 ( t ₁– t ₄) years after subcutaneous specific immunotherapy (SIT). A qualitative reduction ( P =0.05) of the IgE responses to Dp and Der p 2 was observed from t ₁ to t ₄, but a highly statistical significant decrease appeared at t ₃, ( P < 0.01). With regard to Der p 1 IgE values, the immunotherapy induced a significant decrease ( P < 0.01) at t ₃, but not before. In group A, the IgE responses to Der p 1 and Der p 2 were not correlated at t ₀ ( r _s=0.31; P = 0.2l) but were correlated at t₃ ( r _s= 0.78; P=0.001). We also examined sera from 14 adult patients (group B, same SIT schedule as group A) who were without respiratory symptoms at the end of the third year (t₃) of Dp SIT. At this time ( t ₃), there were no significant differences in Der p 1 and Der p 2 IgE levels between group A and group B.