Immunochemical Characterization of Salix alba (S. alba) Pollen Allergens and Evaluation of the Cross-Reactivity Pattern with Common Allergenic Pollen Grains.

BACKGROUND: Pollens, particularly tree and plant pollens, are one of the major causes of allergic respiratory diseases worldwide. Allergy to pollens of different species of Salix trees has been reported in various regions of the world. The most common type of Salix tree in Iran is white willow (Salix alba). OBJECTIVES: This study aimed to identify and determine the immunochemical characteristics of allergenic proteins in S. alba tree pollen extract using SDS-PAGE and IgE- immunoblotting methods. Moreover, the cross-reaction pattern of the specific IgE antibody of S. alba tree pollen proteins with pollen allergens of common allergenic trees, i.e., Populus nigra (P. nigra), Cupressus sempervirens (C. sempervirens), Pinus brutia (P. brutia) and Platanus orientalis (P. orientalis) in the region was investigated. METHODS: The reaction of allergenic proteins in S. alba pollen extract with specific IgE antibodies in patients' sera was investigated using SDS-PAGE and IgE-immunoblotting methods. The cross-reaction of specific IgE antibodies of the proteins present in S. alba pollen extract with pollen allergens of common allergenic trees in the region was investigated using ELISA and immunoblotting inhibition methods. In silico methods such as phylogenetic tree drawing and alignment of amino acid sequences were used to examine the evolutionary relationship and homology structure of common allergenic proteins (Panallergens) responsible for cross reactions. RESULTS: More than 11 protein bands binding to specific IgE antibodies in patients' sera with a molecular weight between 13 and 95 kDa were identified in the S. alba tree pollen extract. ELISA and immunoblotting inhibition results showed that P. nigra extract could inhibit the binding of IgE antibodies to S. alba pollen extract proteins to a greater extent than C. sempervirens, P. brutia, and P. orientalis tree extracts. In silico methods investigated the results of ELISA and immunoblotting inhibition methods. Moreover, a high structural homology and evolutionary relationship were observed between S. alba and P. nigra tree pollen panallergens. CONCLUSION: In this study, it was found that more than 80 % of the sensitive patients who were examined had specific IgE antibodies reacting with the approximately a 15 kDa-protein present in the S. alba pollen extract. Furthermore, the specific IgE-binding proteins found in the pollens of S. alba and P. nigra trees had relative structural homology, and it is likely that if recombinant forms are produced, they can be used for diagnostic and therapeutic purposes for both of the trees.

Immunochemical Characterization of Ligustrum Vulgare (Privet) Pollen Allergens: Study of Common Allergenic Plant in Iran.

Ligustrum vulgare (Privet) pollen proteins are responsible for allergies in susceptible individuals in many regions of the world. This study investigated the immunochemical characterization of Privet pollen extract and the occurrence of skin prick test reactivity to Privet and other allergenic pollen grains in allergic rhinitis patients. All subjects experienced a skin prick test with twenty-two allergen extracts. sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) separated Privet pollen extract, IgE-immunoblotting, and specific ELISA procedures determined the allergenic profile on forty-five Privet allergic patients. A positive allergic reaction to L. vulgare pollen extract was observed in forty-five (31.4%) out of 145 patients. Ten resolved protein fractions were found on SDS-PAGE, ranging from 10 to 80 kDa. IgE-specific antibodies interacted with several allergenic protein bands from Privet-allergic patients in the immunoblotting assay. The most significant interaction was observed in proteins with molecular weights of approximately 15, 18, 43, and 66 kDa. Privet pollen is regarded as a potent allergen composed of IgE-binding constituents. Considering the high allergenicity of Privet pollen grains and since many countries are rich in this plant, identification and production of recombinant forms of common allergens in this species can be used for developing more efficient diagnostic, therapeutic, and preventive approaches.

Molecular and immunochemical characterization of Pop n 2: A new allergen of Populus nigra pollen.

BACKGROUND: Pollen is one of the most common allergens that cause respiratory allergies worldwide. Pollen grains from poplars have been reported as important sources of pollinosis in many countries. OBJECTIVE: The aim of the present study was to determine the molecular and immunochemical characterization of Pop n 2, a novel allergen of Populus nigra (P nigra) pollen extract. METHODS: In this study, the pollen extract of P nigra was analysed by SDS-PAGE, and the allergenic profile was determined by IgE immunoblotting and specific ELISA using the sera of twenty allergic patients. The coding sequence of Pop n 2 was cloned and expressed in the Escherichia coli BL21 (DE3) using plasmid the pET-21b (+). Finally, the expressed recombinant Pop n 2 was purified by affinity chromatography. RESULTS: Pop n 2 belongs to the profilin family with a molecular weight of approximately 14 kDa. Pop n 2 is the most IgE-reactive protein (about 65%) in the P nigra pollen extract. The cDNA sequencing results indicated an open reading frame 396 bp that encodes 131 amino acid residues. The results of ELISA and Immunoblotting assays showed that recombinant Pop n 2 could react with the IgE antibody in patients' sera, like its natural counterpart. CONCLUSION: Our data revealed that Pop n 2 is a significant allergen in the P nigra pollen extract. Moreover, we observed that the recombinant Pop n 2 produced by the pET-21b (+) vector in the E colisystem acts as its natural counterpart.