Type I allergy is an immunoglobulin E (IgE)-mediated hypersensitivity disease affecting more than 25% of the population. Currently, diagnosis of allergy is performed by provocation testing and IgE serology using allergen extracts. This process defines allergen-containing sources but cannot identify the disease-eliciting allergenic molecules. We have applied microarray technology to develop a miniaturized allergy test containing 94 purified allergen molecules that represent the most common allergen sources. The allergen microarray allows the determination and monitoring of allergic patients' IgE reactivity profiles to large numbers of disease-causing allergens by using single measurements and minute amounts of serum. This method may change established practice in allergy diagnosis, prevention, and therapy. In addition, microarrayed antigens may be applied to the diagnosis of autoimmune and infectious diseases.
To cite this article: Brunetto B, Tinghino R, Braschi MC, Antonicelli L, Pini C, Iacovacci P. Characterization and comparison of commercially available mite extracts for in vivo diagnosis. Allergy 2010; 65: 184–190.
Abstract
Background: Assessment of sensitization by allergen‐specific IgE testing and skin prick testing (SPT) are primary tools in routine clinical diagnosis of allergies. To perform a correct diagnosis, it is critical that the allergen reagent used contains an adequate amount of all relevant components. This study aimed at evaluating commercially available mite extracts for in vivo diagnosis from eight manufacturers.
Methods: Eight extracts from Dermatophagoides pteronyssinus and eight from Dermatophagoides farinae were analysed for total protein content by Bradford and for major allergen content by ELISA. SDS‐PAGE, immunoblotting and SPT were also carried out.
Results: The protein amount ranged from 27.7 μg/ml extract to 361.1 μg/ml (D. pteronyssinus) and from 20.3 to 353.0 μg/ml (D. farinae). In regards major allergen concentration, Der p 1 ranged from 9.6 to 36.2 μg/ml, Der f 1 26.5–196.1 μg/ml, mite group 2 0.7–31.7 μg/ml in D. pteronyssinus and 1.3–10.4 μg/ml in D. farinae. SDS‐PAGE experiments showed that some components are poorly represented or absent in extracts from most manufacturers. Similar results were obtained by IgE‐immunoblotting and SPT with 10 mite allergic patients confirmed a broad spectrum of reactivity of the extracts in the same subject.
Conclusions: Immunochemical analysis showed a heterogeneous amount of component/s among mite extracts from different manufacturers. These data were confirmed by in vivo testing, suggesting that, for some of the patient tested, the absence of relevant allergens could strongly affect the diagnosis.
SUMMARYWe report here the expression of b 2 -GPI mRNA by cell types involved in the pathophysiology of the anti-phospholipid syndrome (APS), i.e. endothelial cells as a target of autoantibodies in the APS, astrocytes and neurones involved in APS of the central nervous system (CNS). Lymphocytes were also included in the study, as it has been demonstrated that patients with systemic lupus erythematosusassociated CNS diseases have serum anti-lymphocyte antibodies cross-reacting with brain antigens, and intrathecally synthesized anti-neurone antibodies. Reverse transcriptase-polymerase chain reaction followed by restriction enzyme digestion of the product obtained demonstrated the presence of b 2 -GPI mRNA in all cell types here tested, cultured both in presence and absence of fetal calf serum. In both culture conditions, the same cell types were immunoreactive to an anti-b 2 -GPI MoAb, as determined by indirect immunofluorescence technique. Taken together, these results indicate a direct cell synthesis of b 2 -GPI, suggesting an antigenic function of b 2 -GPI in the APS, including the CNS disease that occurs in this syndrome.
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