The group III allergen from the house dust mite Dermatophagoides pteronyssinus is a trypsin-like enzyme.

Faecally enriched extracts of Dermatophagoides pteronyssinus were shown to contain a trypsin-like enzyme which was allergenic. Chromatofocusing studies revealed the presence of nine major isoforms in D. pteronyssinus, with pI in the range 4 to greater than 8, but only two (range 4-5) in D. farinae. Trypsin isolated from D. pteronyssinus by benzamidine-Sepharose 6B affinity chromatography and gelfiltration was found to be a 31-kDa protein which was enzymatically similar to both invertebrate and vertebrate trypsins. The N-terminal sequence obtained (IVGGEXALAGEXPYQISL) was identical to that reported for the mite allergen Der p III and showed homology with crayfish trypsin and Der f III from D. farinae. Mite trypsin underwent autolysis and the N-terminal sequences of two fragments were found to be ALAGEXPYQI and NNQVXGI respectively. Both showed homology with crayfish trypsin, and the former sequence was identical to residues 7-18 of the native enzyme and Der p III. All isoforms of mite trypsin were showed to be allergenic by radioallergosorbent assay and further studies indicated that the trypsin degradation products were also allergenic. The enzyme was compared with other mite allergens and the rank order of allergenic potency was shown to be: whole mite extract greater than Der p I greater than trypsin. However, all sera from a panel of mite allergic individuals showed IgE reactivity to trypsin, comparable to that seen using whole mite extract and Der p I. These data indicate that mite trypsin is a major allergen corresponding to the previously described allergen, Der p III.     

Bacterial 16S ribosomal DNA in house dust mite cultures

BACKGROUND: Allergen extracts prepared from Dermatophagoides farinae contain significantly more endotoxin than Dermatophagoides pteronyssinus extracts, and extracts from both mite extracts contain more endotoxin than pollen extracts. Attempts to culture bacteria from mite cultures have failed to establish the sources of the endotoxin. OBJECTIVE: To determine the bacterial sources of endotoxin in mite extracts. METHODS: Live mites of both species were obtained from 2 sources, DNA was extracted from the mites, and DNA encoding bacterial 16S ribosomal RNA was amplified by using specific primers. The amount of bacterial DNA in each mite DNA sample was determined by quantitative PCR using an internal standard, and sequence homologies were determined from amplifications performed by using a high-fidelity DNA polymerase. RESULTS: DNA from D farinae appeared to contain between 11-fold and 24-fold more 16S ribosomal gene copies than the genomic DNA from D pteronyssinus (P < or = .003). Sequence analysis indicated the dominant presence of at least 3 phylogenetic clusters of Bartonella species (henselae, quintana, vinsonii, and grahamii), as well as uncharacterized alpha-proteobacteria, from both D farinae and D pteronyssinus. In a few clones, sequences from Escherichia coli, Pseudomonas species, and Acinetobacter species were also identified. CONCLUSION: House dust mite DNA contains evidence of Bartonella and other Gram-negative species. These Gram-negative species are likely to be the sources of the endotoxin found in mite allergenic extracts.     

Enzymatic analyses of house dust mite extracts from Dermatophagoides pteronyssinus and Dermatophagoides farinae (Acari: Pyroglyphidae) during different phases of culture growth.

The majority of clinically important allergens of Dermatophagoides pteronyssinus (Trouessart, 1897) and Dermatophagoides farinae Hughes, 1961 present enzymatic activity. The allergenic enzymes described include cysteine proteases in group 1 allergens, trypsins in group 3, amylases in group 4, and chymotrypsins in group 6. Apart from these, other possibly allergenic enzymes also have been identified. Therefore, enzymatic profiles were studied during the 3 growth periods of the mite population--latency phase, exponential growth phase, and death phase. The activity of 19 different enzymes was analyzed by means of the Api Zym system, a method that has been used to study both mite extracts and other allergenic materials. Our study has demonstrated that the extracts contain a large variety of enzymes. It has been observed that enzymatic activity is caused exclusively by mites because the control carried out on the culture medium was negative for all the enzymes studied. Generally, the levels of diverse enzymatic activity increased with the growth of the culture, and decreased later, in both species. However, proteases are the exception; they maintain a high level of activity during the death phase of the cultured mites. The ratio between trypsin and chymotrypsin activity can be used as an excellent tool for quality control parameters during obtention of allergenic mite extracts.     

Monoclonal immunoassays for major dust mite (Dermatophagoides) allergens, Der p I and Der f I, and quantitative analysis of the allergen content of mite and house dust extracts

Monoclonal, two-site radioimmunoassays (RIAs) were developed to measure allergen Der p I of Dermatophagoides pteronyssinus or Der f I of D. farinae. Microtiter plates coated with monoclonal antibody (Mab) were incubated with mite extract, and bound allergen was detected with a second 125I-labeled Mab of different epitope specificity. The Mab RIAs were very sensitive (nanogram range) and highly specific. D. pteronyssinus extracts with different concentrations of Der p I demonstrated parallel binding curves, whereas a potent D. farinae extract demonstrated less than 5% of the Der p I binding in the same assay. Similar parallel curves were obtained with several D. farinae extracts in the Der f I assay, whereas D. pteronyssinus extract demonstrated little or no binding. The Mab RIAs were compared with an inhibition RIA that measured cross-reacting determinants on both Der p I and Der f I (antigen P1 equivalent [AgP1Eq]). The results demonstrated good quantitative agreement between these assays in commercial mite and house dust extracts (mean difference 1.57 +/- 0.5-fold). Thirty house dust samples with known mite counts, Der p I, and AgP1Eq content were also compared. The summed Mab RIA values for Der p I and Der f I demonstrated a very good correlation with AgP1Eq values (r = 0.86; p less than 0.001) and with assessments of total mite-allergen content by RAST inhibition (n = 21, r = 0.77; p less than 0.001). Furthermore, in samples with more than 10 mites per 100 mg of dust, the Der p I: Der f I ratio closely correlated with the ratio of the two mites counted by microscopy (n = 15, r = 0.89; p less than 0.001). The Mab RIAs can measure allergen levels in mite or dust extracts without the need for purified allergen or affinity-purified antibodies and can readily be standardized. These assays will be useful in epidemiologic studies of allergic asthma, to assess patients' exposure to mite allergens, and the effects of avoidance regimens. Because of the long-term stability and reproducibility of the reagents, Mab-based assays for specific allergens will also play an important role in the standardization of mite and other allergen extracts.     

Comparative detection of mite allergens in house dust of homes in Moscow by enzyme-linked immunosorbent assay and acarologic analysis

The present study revealed that 73% of surveyed apartments in Moscow whose residents included children with the atopic form of bronchial asthma and sensitization to Dermatophagoides pteronyssinus allergens were infested with the pyroglyphid mites D. pteronyssinus and D. farinae. The number of mites in the surveyed apartments varied between 0 and 154 mites/g of dust for D. pteronyssinus and between 0 and 162 mites/g of dust for D. farinae. The levels of mite allergens in these apartments ranged from 0.5 to 165.8 μg/g for Der p I and from 0.3 to 91.3 μg/g of dust for Der f I. The Der p I allergen was found to predominate, and its concentration in one-third of the apartments was more than 10-fold greater than that of Der f I. Correlation between the number of pyroglyphid mites and the concentration of group I allergens was established for both D. pteronyssinus ( r = 0.4932; P < 0.01) and D. farinae (r = 0.6748; P <0.01). In most of the apartments, high and moderate levels of Der I allergens were detected.     

Identification of neryl formate as the airborne aggregation pheromone for the American house dust mite and the European house dust mite (Acari: Epidermoptidae)

The American house dust mite, Dermatophagoidesfarinae Hughes, and European house dust mite, Dermatophagoides pteronyssinus Trouessart, are major pests of medical importance throughout the developed world, causing atopic diseases such as asthma, rhinitis, and atopic dermatitis. Previous studies in our laboratory have shown that the behavioral responses of house dust mites toward volatiles from food sources could be assessed using a Y-tube olfactometer assay. The current study used this Y-tube assay to investigate house dust mite pheromones. A hexane extract of D.farinae, along with fractions of the extract prepared by microscale liquid chromatography over Florisil, were tested for behavioral activity. One of the chromatographic fractions was shown to be significantly attractive (P < 0.05) for D. farinae, compared with a solvent control. Coupled gas chromatography-mass spectrometry analysis of this behaviorally active fraction indicated that neryl or geranyl formate was the major component. Peak enhancement by gas chromatography, using authentic samples of the neryl and geranyl isomers prepared in high purity by chemical synthesis, confirmed the identity of the major peak as neryl formate. In Y-tube assays, male and female D. farinae and D. pteronyssinus both were significantly attracted to synthetic neryl formate at doses of 100 and 10 ng, respectively (P < 0.05). No significant differences were found for D. farinae and D. pteronyssinus when synthetic neryl formate and house dust mite extracts containing natural neryl formate were tested at the same level. Dynamic headspace collection of D. farinae and D. pteronyssinus colonies showed that neryl formate was released as a volatile organic compound by both species. Our study shows that neryl formate is an aggregation pheromone for D. farinae and D. pteronyssinus, and has the potential to be used as part of a novel lure-and-kill system for house dust mite control.     

Faecally derived hydrolytic enzymes from Dermatophagoides pteronyssinus: physicochemical characterisation of potential allergens.

The previous findings that the group I and III mite allergens, and amylase present in mite faeces are hydrolytic enzymes has prompted a study to determine whether this material contains other enzymes which could be allergenic. Thus, spent growth medium devoid of whole Dermatophagoides pteronyssinus mites was shown to contain glucoamylase, lipase and lysozyme in addition to the cysteine protease, serine protease and amylase activities associated with the above allergens, respectively. All of these enzymes are probably associated with mite digestive processes. They were rapidly solubilised, heterogeneous with regard to charge (pI in the range 4-8) and demonstrated maximum biochemical activity in the neutral pH range. Three serine proteases were detected and comprised a chymotrypsin-like, a trypsin-like and an unclassified enzyme with pI of 4.1 and 5.3, 8.5 and 7.1, respectively. Only one cysteine protease was observed, which paralleled immunochemically identified Der p I in a variety of assays. It was shown to cleave at lysyl residues and could be inhibited by the specific cysteine protease inhibitor, E-64. The remaining serine proteases, glucoamylase, lipase and lysozyme represent potential allergens.     

Natural exposure and serum antibodies to house dust mite of mite-allergic children with asthma in Atlanta

Pyroglyphid mites in house dust are important allergens associated with asthma in Europe, but comparable studies of house dust mites in the homes of patients with asthma have not been done in the United States. We examined the distribution of mites and mite allergen in the houses of 20 mite-sensitive children with asthma in Atlanta and measured IgE antibodies to mite allergens in their sera. One or more dust samples from bedding, bedroom floor, television room floor, or television room furniture from 17/20 houses contained greater than 10,000 ng of antigen P1 equivalent per gram of fine dust; amounts ranged from 280 to 230,400 ng/gm. Allergen levels were higher in dust samples from furniture and bedding than from floors. Dust samples obtained from houses in June to September had more mites and mite allergen than those houses sampled in March to April; relative humidity in the room also was higher in June to September. Mite numbers and allergen in floor and furniture samples were correlated with relative humidity in the room and were high when relative humidity was greater than 50%; antigen P1 equivalent was greater than 10,000 ng/gm in 21/39 such samples. Dermatophagoides pteronyssinus was present in all houses and dominant in 11/20. D. farinae was found in 17 houses and was dominant in six. All children studied had high IgE antibody with either D. farinae or D. pteronyssinus RAST; 16 of the 20 children also had IgE to antigen P1. It is likely that the IgE antibody responses in these 20 children with asthma were a direct result of exposure to high levels of mite allergen.(ABSTRACT TRUNCATED AT 250 WORDS)     

利用套式PCR技术鉴别屋尘螨和粉尘螨主要变应原基因及其在尘螨疫苗研制中的应用

通过提取屋尘螨和粉尘螨疫苗研制用的原始种子和生产种子基因组DNA作模板,分别设计Der p1和Der f1各2套内外扩增引物并进行一步法套式PCR,根据扩增出目的片段来检测尘螨变应原Der p1和Der f1基因片段,并以培养基提取物为阴性对照和已通过形态学鉴别为纯屋尘螨、纯粉尘螨的基因组DNA为阳性对照。PCR产物经电泳后切胶回收并克隆到T载体上,进行DNA序列分析并在GenBank中进行同源性比较。结果显示,从屋尘螨和粉尘螨疫苗研制用的原始种子和生产种子分别扩增出含有屋尘螨和粉尘螨主要变应原Der p1和Der f1基因片段,扩增部分的序列与GenBank数据库里相应序列同源性为100%。从阴性对照提取物中未扩增得到目的基因片段,从阳性对照DNA提取物中能扩增得到目的变应原Der p1和Der f1基因片段。本研究首次应用套式PCR技术鉴别了屋尘螨和粉尘螨原始种子和生产种子,为尘螨疫苗研制以及工业化生产提供了一种有效的质量控制手段。     

Enzymatic activity in extracts of allergy-causing astigmatid mites

Many of the previously characterized allergens of house dust mites are known to be proteases, and this enzymatic activity is thought to contribute to their allergenicity. Other astigmatid mites, including stored-product mites and the ectoparasitic itch mite, Sarcoptes scabiei De Geer, are also known to be allergenic, but little or nothing is known about their enzymatic activities. The purpose of this study was to characterize the enzymatic activities present in extracts of the parasitic itch mite and from eight other species of free-living astigmatid mites. Extracts were prepared from one parasitic mite (S. scabiei), five stored-product mites (Chortoglyphus arcuatus (Troupeau), Lepidoglyphus destructor (Schrank), Blomia tropicalis Bronswijk, Cock, Oshima, Tyrophagus putrescentiae (Schrank), and Acarus siro L.), and three house dust mites [Dermatophagoidesfarinae Hughes, Dermatophagoides pteronyssinus (Troussart), and Euroglyphus maynei (Cooreman) ]. ApiZym strips were used to screen for the presence of 19 individual enzyme activities. Digestion of nine other substrates was evaluated by spectrophotometric or electrophoretic methods. All mite extracts exhibited some form of phosphatase, esterase, aminopeptidase, and glycosidase activity, although their substrate specificities varied considerably. Itch mite extract did not possess detectable serine peptidase activity nor was it able to hydrolyze gelatin or casein, whereas all other mite extracts exhibited these activities. Storage mite extracts possessed enzymes capable of degrading the widest range of substrates, whereas itch mite extract had the most limited proteolytic capacity. Extracts of nine species of allergy-causing astigmatid mites contain wide and diverse repertoires of enzymatic activities. These catalytic activities may be important contributors to the induction and manifestation of inflammatory and immune responses to mites in patients.     

Diet Influences Growth Rates and Allergen and Endotoxin Contents of Cultured Dermatophagoides farinae and Dermatophagoides pteronyssinus House Dust Mites

Background: The house dust mites Dermatophagoides farinae and Dermatophagoidespteronyssinus are cultured to obtain material for the production of allergen extracts for research, diagnostic and immunotherapeutic purposes. Methods: We cultured mites on two different diets that supported thriving populations and determined the population growth rates, dynamics of allergen accumulation, and endotoxin concentrations in extracts made from mites harvested from the cultures. Results: D. farinae populations grew faster on a diet of rodent chow/yeast than on an egg/yeast diet but a larger peak population size was achieved on the egg/yeast diet. Diet influenced the dynamics of the production of groups 1 and 2 allergens and the group 1/2 ratios for both species. To population peak, Der f 1 was produced at a faster rate on the chow/yeast diet but greater amounts of Der f 1 were produced by mites grown on the egg/yeast diet. D. pteronyssinus populations grew faster and achieved greater density on the egg/yeast diet compared to the chow/yeast diet. D. pteronyssinus produced more Der p 1 than Der p 2 when grown on chow/yeast while more Der p 2 than Der p 1 was produced on egg/yeast. Endotoxin concentrations in extracts made from whole cultures for both species at maximum population density were very different in the two diets. Washing the mites resulted in the loss of up to 88% of the allergen. Conclusion: Mite-culturing diet directly effects population growth, the dynamics of allergen accumulation, the group 1/2 allergen ratio and the endotoxin contents in extracts of cultured house dust mites.     

Sensitization and exposure to house dust and storage mites in high-altitude areas of ecuador.

BACKGROUND: Few studies have addressed exposure and sensitization to mite allergens in Andean countries. OBJECTIVES: To identify the main mite species in 3 locations at different altitudes in Ecuador and to verify skin test reactivity to various mite species in allergic individuals in Quito, Ecuador. METHODS: Mattress dust samples were collected in Quito (2,800 m above sea level), Cuenca (2,500 m above sea level), and Guayaquil (sea level). Mite species present in the samples were isolated, identified, and counted. Der p 1 and Der f 1 levels were measured using monoclonal antibody-based enzyme immunoassays. Four hundred thirty-five patients in Quito diagnosed as having allergic rhinitis or asthma underwent skin testing with commercial extracts of Dermatophagoides pteronyssinus, Dermatophagoides farinae, Blomia tropicalis, Tyrophagus putrescentiae, and Lepidoglyphus destructor. In addition, Glycyphagus domesticus, Acarus siro, and Aleuroglyphus ovatus were tested in 362, 262, and 279 patients, respectively. RESULTS: Twenty-one mite species were identified. Large populations of mites were detected above 2,500 m of altitude. All the dust samples contained detectable levels of Der p 1 or Der f 1. Positive skin prick test reactions to D. pteronyssinus, D. farinae, B. tropicalis, L. destructor, T. putrescentiae, A. ovatus, A. siro, and G. domesticus were obtained in 60.9%, 56.8%, 17.0%, 19.3%, 10.6%, 15.8%, 8.8%, and 11.0% of the patients, respectively. CONCLUSIONS: Most analyzed mattresses contained several species of mites. Mite allergen levels were high. This study confirms the importance of house dust and storage mite allergens in Ecuador in areas above 2,500 m of altitude, where humidity remains high year round.     

High-molecular-weight allergens of the house dust mite: an apolipophorin-like cDNA has sequence identity with the major M-177 allergen and the IgE-binding peptide fragments Mag1 and Mag3

BACKGROUND: A 349-residue recombinant polypeptide of Dermatophagoides farinae, Mag 3, has been shown to represent part of a larger 177-kD (M-177) allergen with very high IgE-binding activity. METHODS: Cloning and sequencing of cDNA from the house dust mites Dermatophagoides pteronyssinus and Euroglyphus maynei was used to characterise the polypeptide containing the Mag 3 sequence. RESULTS: cDNA clones containing the complete sequence of the E. maynei homologue of the M-177 allergen were isolated and analysed. The translation contained not only an amino acid sequence with 90% identity to the 349-residue Mag-3 fragment but also a further sequence with 90% identity to another IgE-binding recombinant D. farinae polypeptide designated Mag 1. The complete sequence encoded a mature polypeptide of 1,650 residues and a molecular mass of 189.5 kD. cDNA clones from D. pteronyssinus also encoded sequences equivalent to the Mag 1 and 3 polypeptides. The M-177 sequence showed strong similarity to the lipid transport apolipophorins found in insect lipophorins. CONCLUSIONS: cDNA sequence data show that the D. pteronyssinus and E. maynei homologues of the M-177 high-molecular-weight D. farinae allergen contain sequences equivalent to both the Mag 1 and Mag 3 recombinant IgE-binding fragments. The N-terminal sequence of the full-length 1,650 amino-acid allergen showed strong similarity to the insect apolipophorins which are poorly soluble in aqueous extracts and exist in the lipid transport particles in haemolymph. It is proposed that presentation in lipid particles could be a factor which enhances the immunogenicity of this group of allergens.     

House dust mite sensitivity: a comparison of immune response with exposure to four species of Pyroglyphidae

Twenty-five atopic children under 11 years of age were studied, using skin and RAST tests, for their specific IgE response to four species of pyroglyphid house dust mites, Dermatophagoides pteronyssinus, D. farinae, D. microceras and Euroglyphus maynei. All of the children were sensitive to D. pteronyssinus, 20 (80%) of these children were also sensitive to D. farinae and D. microceras, and 16 of the latter (64%) were also sensitive to E. maynei. Dust samples from various sites in the homes of the children revealed D. pteronyssinus in all homes studied but no D. farinae or D. microceras. E. maynei, although identified, was not present in significant numbers in any site. A control group of 20 atopic children of similar age who were not sensitive to house dust mite allergens had a similar exposure to the four mite species. These results suggest that factors in addition to mite exposure are important in the development of specific IgE responses to house dust mites.     

Production and proteomic characterization of pharmaceutical-grade Dermatophagoides pteronyssinus and Dermatophagoides farinae extracts for allergy vaccines

BACKGROUND: House dust mites (HDM) such as Dermatophagoides pteronyssinus and Dermatophagoides farinae represent a major cause of type 1 allergies worldwide. Hence large quantities of well-characterized HDM extracts are needed to prepare pharmaceutical-grade allergy vaccines. To this aim, the present study was undertaken to define optimal conditions for large-scale cultures. METHODS: D. pteronyssinus and D. farinae were grown on different media combining various proportions of wheat germ, yeast and synthetic amino acids (the latter resembling the composition of the human stratum corneum). Extracts thus obtained were analyzed for their total allergenic activity, as well as major allergen and protein contents, using immunosorbent assays, HPLC, immunoblotting, two-dimensional electrophoresis and peptide mass fingerprinting. RESULTS: An optimal culture medium (Stalmite APF) based on wheat germ, yeast and amino acids in defined proportion (42, 42 and 15% w/w, respectively) was selected to grow various HDM species with high yields. A detailed proteomic analysis revealed that D. pteronyssinus extracts generated under such conditions did not contain allergens originating from culture medium components and that major prevalent HDM allergens (i.e. groups 1, 2, 7, 10, 13 and 20) are found among the most abundant proteins in the D. pteronyssinus extract. Semiquantitative dot-blot assays confirmed the presence of Der p 3-10 as well as Der p 13 and 14 allergens within the extracts. CONCLUSIONS: We developed a well-defined medium allowing to grow various HDM species at an industrial scale in a highly reproducible manner. Extracts from mites produced under such pharmaceutical conditions contain all the relevant allergens for desensitization purposes and in vivo diagnosis.     

Treatment of allergy to house dust with pyridine-extracted alum-precipitated extracts of the house dust mite.

Two preliminary trials in which patients with respiratory allergy to the house dust mite were treated with pyridine-extracted alum-precipitated extracts of house dust mites are described. In the first, using an extract of D. farinae, twenty-two of twenty-eight asthmatic patients improved. In the second, using an extract of D. pteronyssinus, fifteen of eighteen asthmatic patients improved.     

Sensitization to Dermatophagoides siboney, Blomia tropicalis, and other domestic mites in asthmatic patients

Mite species adapted to warm, humid climates are commonly found in house dust in the tropics. In Cuba, Dermatophagoides pteronyssinus, D. siboney , and Blomia tropicalis are the most common and abundant mite species in house dust. To investigate the pattern of Sensitization of Cuban asthmatic patients to common mite species, we skin-prick-tested (SPT) 148 patients with a clinical history of asthma and possible mite allergy, and determined specific IgE antibodies against mite allergens ( D. pteronyssinus, D. farinae, D. siboney, B. tropicalis, Acarus siro, Lepidoglyphus destructor, Tyrophagus putrescentiae , and Glycyphagus domesticus ). The prevalence of positive SPT was high to D. siboney (88%), D. pteronyssinus (87%), A. siro (85%), B. tropicalis (85%), and D. farinae (83%). The largest skin reactions were obtained with D. siboney and B. tropicalis extracts. The skin test response to the D. siboney extract correlated to those of D. farinae, D. pteronyssinus, B. tropicalis , and A. siro. The highest IgE levels were found to Dermatophagoides species and B. tropicalis. IgE to D. siboney and B. tropicalis were found in 97% and 96% of the patients, respectively. The prevalence of specific IgE to the other mites studied varied from 46 to 65%. D. siboney and B. tropicalis are important sensitizers among asthmatic patients in Cuba.     

Dermatophagoides farinae (Der f 1) and Dermatophagoides pteronyssinus (Der p 1) allergen exposure among subjects living in Uberlândia, Brazil

BACKGROUND: The role of mite allergen exposure in sensitization and development of asthma has been widely recognized. Previous studies have shown that Dermatophagoides pteronyssinus and Blomia tropicalis were the most prevalent house dust mites in Brazil, while D. farinae was rarely found. The aim of this study was to measure Der f 1 and Der p 1 allergen levels in Brazilian asthmatics' and controls' homes. METHODS: Sixty-four houses (32 asthmatic, 32 control) were visited for dust sampling from five sites. Der f 1 and Der p 1 levels were measured by two-site monoclonal-antibody-based ELISAs. RESULTS: The highest levels of Der f 1 and Der p 1 allergens were found in bedding samples from both asthmatics' and controls' homes. However, the geometric mean of Der f 1 levels (15.8 microgram/g of dust) was significantly higher than for Der p 1 (8.2 microgram/g of dust) in these samples. In addition, allergen levels >/=10 microgram/g of dust were found in 60-80% of the samples for Der f 1 and about 50% for Der p 1. CONCLUSIONS: These results indicate that high levels of Der f 1 allergen are present in both asthmatics' and controls' homes, in contrast to previously reported data. Therefore, studies on exposure to mites should be performed in different cities, seasons and times, since the mite fauna might be subject to variations. Knowledge of the mite fauna will certainly improve the means of investigating the association between allergen exposure and sensitization, allowing to establish the inclusion of new mite extracts in inhalant skin test sets, and even to detect monosensitized patients with respiratory allergy.     

Does guanine concentration in house-dust samples reflect house-dust mite exposure?

Using specific ELISA extracts, we analyzed samples of pure mite cultures and house dust for group I allergens from Dermatophagoides pteronyssinus , D. farinae , and D. microceras . Correspondingly, we measured the concentration of guanine and xanthine by rigorous chemical methods. The pure mite cultures were used to correiate allergen levels with guanine levels. The house-dust samples one-sidedly contained more guanine than expected. This suggests that considerable amounts of guanine originate from non-house-dust mite sources and implies that guanine quantifications are of limited clinical value.     

Immunoblot multi-allergen inhibition studies of allergenic cross-reactivity of the dust mites Lepidoglyphus destructor and Dermatophagoides pteronyssinus

The allergenic similarity of the pyroglyphid mite D. pteronyssinus and the glycyphagid mite L. destructor was investigated with a new immunoblotting inhibition technique allowing simultaneous comparison of several allergens. Extracts of D. pteronyssinus and L. destructor were separated by SDS-PAGE and electroblotted to nitrocellulose (NC). A serum pool containing IgE specific to the major allergens in both mites was mixed with serially diluted extracts of D. pteronyssinus and L. destructor and incubated with the mite allergens of NC. The inhibition of the IgE binding to NC was evaluated by densitometric scanning and percentage inhibition was calculated. The IgE antibodies to the 25-kD component in D. pteronyssinus, were inhibited to the same degree by extracts of D. pteronyssinus and L. destructor. Another major allergen component in D. pteronyssinus (16 kD) was also inhibited by L. destructor extract but to a lesser degree: 400 times more of the heterologous than of the homologous extract was needed for 50% inhibition. To produce 50% of heterologous inhibition of the two major allergen components at 15 and 53 kD of L. destructor, 2000 and 10,000 times more respectively, of D. pteronyssinus than of L. destructor extract were needed. Two minor allergen components of L. destructor showed some cross-reactivity with D. pteronyssinus. However, L. destructor was a stronger inhibitor of D. pteronyssinus than vice versa, probably because the sera were obtained from persons more sensitized to L. destructor than to D. pteronyssinus.