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The development of rapid analysis of human serum for the presence of allergen-specific Immunoglobulin E (IgE) is currently important. Consequently, we developed two types of three-dimensional (3D) protein biochips. The first one is a 3D hydrogel biochip containing hydrogel droplets with protein molecules (allergens, immunoglobulins and others). These droplets are disposed on elements consisting of short polymer brushes grafting from a surface of polybutylene terephthalate polymer. The immobilization of proteins was induced by short-wave ultraviolet (UV) radiation. On such a biochip, the kinetics of allergen–sIgE complex formation reached 60% of saturation for 6 h. Also, we developed a 3D brush microchip containing on the surface of a polyethylene terephthalate polymer the brush elements with protein molecules covalently immobilized by opening oxirane cycles by amino and thiol nucleophilic groups contained in proteins. In the case of the 3D brush microchip, the kinetics of allergen–sIgE complex formation reached 100% of saturation for 3 h, and fluorescent signals were 2–3 times higher than those of the 3D hydrogel biochip for some allergens. Thus, the comparative analysis revealed that 3D brush biochips are more useful for further studies of protein–protein interaction than 3D hydrogel ones.
The development of rapid analysis of human serum for the presence of allergen-specific Immunoglobulin E (IgE) is currently important. Consequently, we developed two types of three-dimensional (3D) protein biochips. The first one is a 3D hydrogel biochip containing hydrogel droplets with protein molecules (allergens, immunoglobulins and others). These droplets are disposed on elements consisting of short polymer brushes grafting from a surface of polybutylene terephthalate polymer. The immobilization of proteins was induced by short-wave ultraviolet (UV) radiation. On such a biochip, the kinetics of allergen–sIgE complex formation reached 60% of saturation for 6 h. Also, we developed a 3D brush microchip containing on the surface of a polyethylene terephthalate polymer the brush elements with protein molecules covalently immobilized by opening oxirane cycles by amino and thiol nucleophilic groups contained in proteins. In the case of the 3D brush microchip, the kinetics of allergen–sIgE complex formation reached 100% of saturation for 3 h, and fluorescent signals were 2–3 times higher than those of the 3D hydrogel biochip for some allergens. Thus, the comparative analysis revealed that 3D brush biochips are more useful for further studies of protein–protein interaction than 3D hydrogel ones.
BACKGROUND: Due to the widespread occurrence of respiratory allergic diseases, it is of particular interest to study the regional spectrum of sensitization of the population using a modern method-molecular allergodiagnostics. OBJECTIVE: to determine the spectrum of sensitization in children with allergies living in the Rostov region by the method of molecular allergodiagnostics. METHODS: An open multicenter prospective controlled non-randomized study included 67 children with respiratory allergic diseases living in the Rostov region. The patients were examined using the Allergy Explorer 2 (ALEX2) allergy chip. Statistical processing of the results was carried out using the Microsoft Office Excel 2003 and Statistica 12.0 for Windows software package. RESULTS: According to the conducted studies, it was found that the most common sensitization in children of the Rostov region is caused by weed pollen, pet allergens, pollen of trees, cereals, fruits, mold and yeast fungi, class 4 sensitization was also noted to uteroglobin and lipocalin. Molecular analysis showed that an allergic reaction to ragweed pollen prevailed among pollen allergens - the allergocomponent Amb a1 caused sensitization in 27 (40.3%) children, and nettle (Urt d) - 23 (34.33%) of the examined. The analysis of the allergen components of tree pollen showed that the most common positive reaction was to date palm (22.39%), cryptomeria japonica (19.4%), while only 6 (8.96%) patients had a "true" allergy to birch pollen (positive component Bet v 1). If we talk about sensitization to house dust mites and mold allergens, it should be noted that the molecules Alternaria alternata (20.89%), Blomia tropicalis (23.88%) and Glycyphagus domesticus (23.88%) prevailed in the spectrum of this group of allergens. CONCLUSION. In the course of the study, it was found that most often in children of the Rostov region with respiratory allergic diseases, sensitization to weed pollen was determined. This study allowed not only to determine the class of sensitization, but also to identify the allergen components that play a leading role in the development of pathology.
Food allergies are one of the most common variants of allergic diseases in children. Nowadays it is necessary to apply the most sensitive, specific and standardized methods for the detection of allergensin theoretical allergology and clinical practice.For this purpose, the detection of specific immunoglobulins of class E (sIgE) in blood serum is most often used, however the basophil activation test (BAT) deserves close attention due to its functional methodology based on its maximum proximity to the pathogenetic mechanisms of immune reactions.In order to compare the results of detecting sensitization to food allergens in BAT and the method of detecting specific IgE 76 patients aged 1-16 years receiving outpatient treatment with a pediatrician or an allergist-immunologist for gastrointestinal pathology were examined. During the analysis two subgroups were identified: patients with a burdened allergic anamnesis (36 people) and patients without clear anamnestic data on the presence of food allergies (40 people). The activation of basophils to allergens to cow’s milk, egg, wheat and soybeans proteins was evaluated by flow cytometry using the Allergenicitykit test system (‘BeckmanCoulter’), specific IgE was evaluated by using the AllergoIFE-specific IgE kit (‘Alkor Bio’).The results showed that the most common significant allergen of food allergy in the general group was cow’s milk proteins, at the same time the number of positive BAT results exceeded the number of cases of allergy detection by the method of determining specific IgE. Specific IgE (f2) was detected in 7 people, sIgE (f77) was detected in 3 patients, and sIgE (f78) – in 5 children in the subgroup of patients with suspected allergy to cow’s milk proteins. Using BAT obtained a positive result in 26, 21 and 13 cases, respectively. In a subgroup of children with gastrointestinal pathology without clear anamnestic data on food sensitization specific IgE(f2) was positive in 1 patient, sIgE (f77) – in 1 person, sIgE (f78) was detected in 4 children. BAT-positive result was obtained in 25, 24, 11 cases, respectively.The results demonstrate the advantage of BAT in comparison with sIgE detection. The inclusion of BAT in the examination plan for children with incomplete effectiveness of standard treatment of gastrointestinal diseases will increase the detection of allergopathologies.
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