CHARACTERIZATION OF rBlo t 13, RECOMBINANT ALLERGEN FROM Blomia tropicalis USING MONOCLONAL ANTIBODIES.

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Bibliographic Details
Title:	CHARACTERIZATION OF rBlo t 13, RECOMBINANT ALLERGEN FROM Blomia tropicalis USING MONOCLONAL ANTIBODIES.
Authors:	Labrada, M.¹, Uyema, K.¹, Sewer, M.¹, Labrada, A.¹ labrada@biocen.colombus.cu, González, M.¹, Puerta, L.², Caraballo, L.²
Source:	Revista VacciMonitor (Vacunología y Temas Afines). Oct2002, Vol. 11 Issue 4, p1. 1p.
Subjects:	ALLERGENS, MONOCLONAL antibodies, MITES, RECOMBINANT proteins, IMMUNOGLOBULIN E, HYBRIDOMAS, DERMATOPHAGOIDES
Abstract:	Blo t13 is a 17 kD allergen of the domestic mite Blomia tropicalis (BT), that have been cloned and expressed as a recombinant protein in E. coli and Pichia pastoris. It shows high IgE reactivity to sera of allergic patients. The biological function corresponds to fatty acid binding protein family; however, the immunological properties are not well studied yet. This work describes the production of monoclonal antibodies (MAbs) specific to Blo t13, and their use in the antigenic characterization of this allergen. Balb/c mice were immunized with rBlo t13 expressed in P.pastoris. Hybridoma screening was performed using a direct ELISA with solid-phase bound recombinant allergen. Hybrids producing anti-rBlo t13 specific antibodies were cloned. MAb specificity was tested by immunoblotting. Topography of the binding site and MAb binding to recombinant and native allergen were studied by different ELISA assays. Two hybridomas producing MAbs (IgG1 isotype) against Blo t13 were generated. MAbs specifically recognized the 17 kD recombinant protein in the immunoblotting experiment. Both antibodies recognized the same or closed epitopes on the rBlo t13 molecule, as was determined by mutual inhibition in ELISA experiments. The results suggested that this epitope should be repeated on the molecule. The binding of MAbs to rBlo t13 was also inhibited by BT allergenic extract, confirming the immunological identity of the native and recombinant molecules. A high degree of inhibition was also obtained using a Dermatophagoides siboney extract, suggesting the presence of a Blo t13 analog in this mite specie. [ABSTRACT FROM AUTHOR]
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Database:	MedicLatina

Description
Abstract:	Blo t13 is a 17 kD allergen of the domestic mite Blomia tropicalis (BT), that have been cloned and expressed as a recombinant protein in E. coli and Pichia pastoris. It shows high IgE reactivity to sera of allergic patients. The biological function corresponds to fatty acid binding protein family; however, the immunological properties are not well studied yet. This work describes the production of monoclonal antibodies (MAbs) specific to Blo t13, and their use in the antigenic characterization of this allergen. Balb/c mice were immunized with rBlo t13 expressed in P.pastoris. Hybridoma screening was performed using a direct ELISA with solid-phase bound recombinant allergen. Hybrids producing anti-rBlo t13 specific antibodies were cloned. MAb specificity was tested by immunoblotting. Topography of the binding site and MAb binding to recombinant and native allergen were studied by different ELISA assays. Two hybridomas producing MAbs (IgG1 isotype) against Blo t13 were generated. MAbs specifically recognized the 17 kD recombinant protein in the immunoblotting experiment. Both antibodies recognized the same or closed epitopes on the rBlo t13 molecule, as was determined by mutual inhibition in ELISA experiments. The results suggested that this epitope should be repeated on the molecule. The binding of MAbs to rBlo t13 was also inhibited by BT allergenic extract, confirming the immunological identity of the native and recombinant molecules. A high degree of inhibition was also obtained using a Dermatophagoides siboney extract, suggesting the presence of a Blo t13 analog in this mite specie. [ABSTRACT FROM AUTHOR]
ISSN:	1025028X