Rosetta Ferrara

Cross-sectional survey on immunoglobulin E reactivity in 23 077 subjects using an allergenic molecule-based microarray detection system

Background The availability of allergenic molecules and high‐throughput microtechnologies allow the collection of a large number of IgE results at the same time in a single test. This can be carried out applying the test in the routine diagnostic work‐up.

Identification of Der p 23, a Peritrophin-like Protein, as a New Major Dermatophagoides pteronyssinus Allergen Associated with the Peritrophic Matrix of Mite Fecal Pellets

The house dust mite (HDM) Dermatophagoides pteronyssinus is one of most important allergen sources and a major elicitor of allergic asthma. We screened a D. pteronyssinus expression cDNA library with IgE Abs from HDM allergic patients. A cDNA coding for a new major allergen was isolated, which showed sequence homology to peritrophins, which contain chitin-binding domains and are part of the peritrophic matrix lining the gut of arthropods. The mature Der p 23 allergen was expressed in Escherichia coli as an 8-kDa protein without its hydrophobic leader sequence and purified to homogeneity. It reacted with IgE Abs from 74% of D. pteronyssinus allergic patients (n = 347) at levels comparable to the two major HDM allergens, Der p 1 and Der p 2. Thus, Der p 23 represents a new major D. pteronyssinus allergen. Furthermore, rDer p 23 exhibited high allergenic activity as demonstrated by upregulation of CD203c expression on basophils from D. pteronyssinus allergic patients. Immunogold electron microscopy localized the allergen in the peritrophic matrix lining the midgut of D. pteronyssinus as well as on the surface of the fecal pellets. Thus, we identified a new major D. pteronyssinus allergen as peritrophin-like protein. The high allergenic activity of Der p 23 and its frequent recognition as respiratory allergen may be explained by the fact that it becomes airborne and respirable through its association with mite feces. Der p 23 may be an essential component for diagnosis and specific immunotherapy of HDM allergy.

Varying Allergen Composition and Content Affects the in vivo Allergenic Activity of Commercial Dermatophagoides pteronyssinus Extracts

Background: Diagnosis and immunotherapy of house-dust mite (HDM) allergy is still based on natural allergen extracts. The aim of this study was to analyze commercially available Dermatophagoides pteronyssinus extracts from different manufacturers regarding allergen composition and content and whether variations may affect their allergenic activity. Methods: Antibodies specific for several D. pteronyssinus allergens (Der p 1, 2, 5, 7, 10 and 21) were used to analyze extracts from 10 different manufacturers by immunoblotting. Sandwich ELISAs were used to quantify Der p 1 and Der p 2 in the extracts. Mite-allergic patients (n = 45) were skin-tested with the extracts and tested for immunoglobulin E (IgE) reactivity to a panel of 10 mite allergens (Der p 1, 2, 4, 5, 7, 8, 10, 14, 20 and 21) by dot blot. Results: Only Der p 1 and Der p 2 were detected in all extracts but their concentrations and ratios showed high variability (Der p 1: 6.0–40.8 µg ml–1; Der p 2: 1.7–45.0 µg ml–1). At least 1 out of 4 allergens (i.e. Der p 5, 7, 10 and 21) was not detected in 8 of the studied extracts. Mite-allergic subjects showed different IgE reactivity profiles to the individual mite allergens, the extracts showed different allergenic activity in skin-prick tests and false-negative results. Conclusions: Commercially available D. pteronyssinus extracts lack important allergens, show great variability regarding allergen composition and content and some gave false-negative diagnostic test results in certain patients.

Ovomucoid (Gal d 1) specific IgE detected by microarray system predict tolerability to boiled hen's egg and an increased risk to progress to multiple environmental allergen sensitisation

Egg allergy is a very common finding in early childhood. Detecting hens egg (HE) allergy outgrowing and reintroduction of food containing egg is a task for the allergist.

Der p 11 Is a Major Allergen for House Dust Mite-Allergic Patients Suffering from Atopic Dermatitis

House dust mites (HDMs) belong to the most potent indoor allergen sources worldwide and are associated with allergic manifestations in the respiratory tract and the skin. Here we studied the importance of the high-molecular-weight group 11 allergen from Dermatophagoides pteronyssinus (Der p 11) in HDM allergy. Sequence analysis showed that Der p 11 has high homology to paramyosins from mites, ticks, and other invertebrates. A synthetic gene coding for Der p 11 was expressed in Escherichia coli and rDer p 11 purified to homogeneity as folded, alpha-helical protein as determined by circular dichroism spectroscopy. Using antibodies raised against rDer p 11 and immunogold electron microscopy, the allergen was localized in the muscle beneath the skin of mite bodies but not in feces. IgE reactivity of rDer p 11 was tested with sera from HDM-allergic patients from Europe and Africa in radioallergosorbent test-based dot-blot assays. Interestingly, we found that Der p 11 is a major allergen for patients suffering from atopic dermatitis (AD), whereas it is only a minor allergen for patients suffering from respiratory forms of HDM allergy. Thus, rDer p 11 might be a useful serological marker allergen for the identification of a subgroup of HDM-allergic patients suffering from HDM-associated AD.

Carrier‐bound Alt a 1 peptides without allergenic activity for vaccination against Alternaria alternata allergy

The mould Alternaria alternata is a major elicitor of allergic asthma. Diagnosis and specific immunotherapy (SIT) of Alternaria allergy are often limited by the insufficient quality of natural mould extracts.

HIV-specific lymphoproliferative responses in asymptomatic HIV-infected individuals

In vitro lymphoproliferative responses to HIV‐1 recombinant antigens (gp160, p24, and Rev protein) were studied in 83 patients with asymptomatic HIV‐1 infection (CDC groups 11 and III) and circulating CD4 lymphocyte numbers > 400/mm3. Significant response to at least one of the three antigens was detected in 52.4% of the subjects, but the responses were weak, and concordance of the response to the three antigens was rare, the frequency of individuals responding to each antigen not exceeding 22.4%. Increasing frequencies of response were observed when recall antigens (tetanus toxoid and Candida albicans glycomannoprotein) (65.5%) and anti‐CD3 MoAb (76.6%) were used as stimuli. Although a significant association between lymphocyte response to p24, but not gp160, and steadiness of CD4 lymphocyte numbers before the assay was observed, no predictive value for lack of CD4 cell decrease was confirmed for either antigen, and fluctuation of the responses to HIV antigens was seen during subsequent follow up. The panel of T cell assays used could be regarded as appropriate for monitoring both HIV‐specific responses and T lymphocyte function during immunotherapy with soluble HIV antigens.

Peamaclein – A new peach allergenic protein: similarities, differences and misleading features compared to Pru p 3

Among the peach‐derived allergens which are already known, the lipid transfer protein (Pru p 3) seems to be the one to exert severe allergic reactions.

Tolerability of a Fully Maturated Cheese in Cow’s Milk Allergic Children: Biochemical, Immunochemical, and Clinical Aspects

Background From patients’ reports and our preliminary observations, a fully maturated cheese (Parmigiano-Reggiano; PR) seems to be well tolerated by a subset of cow’s milk (CM) allergic patients. Objective and Methods To biochemically and immunologically characterize PR samples at different maturation stage and to verify PR tolerability in CM allergic children. Seventy patients, with suspected CM allergy, were enrolled. IgE to CM, α-lactalbumin (ALA), β-lactoglobulin (BLG) and caseins (CAS) were tested using ImmunoCAP, ISAC103 and skin prick test. Patients underwent a double-blind, placebo-controlled food challenge with CM, and an open food challenge with 36 months-maturated PR. Extracts obtained from PR samples were biochemically analyzed in order to determine protein and peptide contents. Pepsin and trypsin-chymotrypsin-pepsin simulated digestions were applied to PR extracts. Each PR extract was investigated by IgE Single Point Highest Inhibition Achievable assay (SPHIAa). The efficiency analysis was carried out using CM and PR oral challenges as gold standards. Results The IgE binding to milk allergens was 100% inhibited by almost all PR preparations; the only difference was for CAS, mainly αS1-CAS. Sixteen patients sensitized to CM tolerated both CM and PR; 29 patients tolerated PR only; 21 patients, reacted to both CM and PR, whereas 4 patients reactive to CM refused to ingest PR. ROC analysis showed that the absence of IgE to BLG measured by ISAC could be a good marker of PR tolerance. The SPHIAa using digested PR preparations showed a marked effect on IgE binding to CAS and almost none on ALA and BLG. Conclusions 58% of patients clinically reactive to CM tolerated fully maturated PR. The preliminary digestion of CAS induced by PR maturation process, facilitating a further loss of allergenic reactivity during gut digestion, might explain the tolerance. This hypothesis seems to work when no IgE sensitization to ISAC BLG is detected.

Wheat IgE profiling and wheat IgE levels in bakers with allergic occupational phenotypes

Objectives To characterise occupational wheat allergic phenotypes (rhino-conjunctivitis, asthma and dermatitis) and immunoglobulin (IgE) sensitisation to particular wheat allergens in bakers. Methods We conducted clinical and immunological evaluations of 81 consecutive bakers reporting occupational symptoms using commercial tests (skin prick test (SPT), specific IgE, ISAC microarray) and six additional dot-blotted wheat allergens (Tri a 39, Tri a Trx, Tri a GST, Tri a 32, Tri a 12, Tri a DH). Results Wheat SPT resulted positive in 29 bakers and was associated with work-related asthma (p<0.01). Wheat IgE was detected in 51 workers and was associated with work-related asthma (p<0.01) and rhino-conjunctivitis (p<0.05). ISAC Tri a 30 was positive in three workers and was associated with work-related dermatitis (p<0.05). Wheat dot-blotted allergens were positive in 22 bakers. Tri a 32 and Tri a GST were positive in 13 and three bakers, respectively, and both were associated with work-related dermatitis (p<0.05). This association increased (p<0.01) when Tri a 32, Tri a GST and Tri a 30 were analysed together (p<0.01). Wheat IgE levels were associated with work-related dermatitis (p<0.01). Conclusions Wheat IgE levels and wheat microarrayed allergens may be associated with some occupational allergic phenotypes. The extension of the panel of wheat allergens may be promising for discriminating the clinical manifestations of bakers allergy.