Monoclonal Antibodies against Der s 1, a Major Allergen of <i>Dermatophagoides siboney</i>

Abstract

Six stable clones secreting murine monoclonal antibodies (Mabs) against Der s 1 were obtained. The binding of Mabs showed cross-reactivity with Dermatophagoides farinae, as determined by enzyme-linked immunosorbent assay (ELISA). In a Western blot assay, antibodies reacted with a 24-kD protein considered to represent the major allergen Der s 1. The repertoire of antigenic sites on Der s 1 was studied using a panel of Mabs. Epitope specificity of the Mabs was determined by both competitive inhibition and sandwich ELISA assays. The results defined six different, non-overlapping and non-repeated antigenic sites on the allergen molecule. Der s 1 allergen from Dermatophagoides siboney extracts was purified by Mab affinity chromatography, this procedure gave 43% recovery of >90% pure allergen. The purified allergen had capacity to bind specific human IgE and demonstrated an allergenic activity of up to 77% of total D. siboney extract. An Mab-ELISA was developed using Mabs directed against different epitopes on Der s 1. This assay could detect up to 1 ng/ml of Der s 1 and Der f 1 in allergen preparations.

Allergen Micro-Bead Array for IgE Detection: A Feasibility Study Using Allergenic Molecules Tested on a Flexible Multiplex Flow Cytometric Immunoassay

“…For the first set of experiments, lyophilized purified natural (n) Der s 1, a group 1 mite allergen from Dermatophagoides siboney , was provided by Biocen [17] (Havana, Cuba), nPen m 1, a tropomyosin from a shrimp species ( Penaeus monodon ) was purchased from BIAL-Aristegui (Bilbao, Spain), and nPru p 3, a lipid transfer protein (LTP), was purified from peaches ( Prunus persica ) as previously reported [18]. nPru p 3 preparation immobilized on ISAC was not the same used for micro-bead conjugation.…”

“…To detect nDer s 1 onto the micro-beads, two different MoAbs were used, an IgG1 isotype (4E10/E10) and an IgG2b isotype (5F7/H8F2) (Biocen) [17]. For detecting nPen m 1 conjugation to the micro-beads an anti-shrimp tropomyosin PoAb was used (BIAL-Aristegui).…”

Immunogenicity of a novel anti-allergic vaccine based on house dust mite purified allergens and a combination adjuvant in a murine prophylactic model

“…For this, an MPC Aerosol Medication Nebulizer (Braintree Scientific, Braintree, MA) was connected to a plastic chamber where the mice were kept during the challenge ( 24 ). The allergen solution used for nebulization was the same allergen mix DS1/DS2 (500 µg/ml Der s 1 ( 25 ), 100 µg Der s 2 (as measured by specific Der f 2 ELISA kit; Indoor Biotechnologies, Charlottesville, VA)). After taking into account, both starting concentration and chamber volume, the actual allergen concentration in air was calculated as 105 µg/L Der s 1 and 21 µg/L Der s 2.…”