Amoxicillin (AX)is nowadays the β-lactam that more frequently induces immediate allergic reactions. Nevertheless, diagnosis of AX allergy is occasionally challenging due to risky in vivo tests and non-optimal sensitivity of in vitro tests. AX requires protein haptenation to form multivalent conjugates with increased size to be immunogenic. Knowing adduct structural features for promoting effector cells activationwould help to improve in vitro tests. We aimed to identify the optimal structural requirement in specific cellular degranulation to AX using well-precised nanoarchitecturesof different lengths.
We constructed eightBidendron Antigens (BiAns) based on polyethylene glycol(PEG) linkers of different lengths (600-12000 Da), end-coupled with PolyAmidoAmine dendrons that were terminally multi-functionalized with amoxicilloyl (AXO).In vitro IgE recognition was studied by competitive RadioAllergoSorbentTest (RAST) and antibody-nanoarchitecture complexes by transmission electron microscopy (TEM). Their allergenic activity was evaluated using bone marrow-derived mast cells (MCs) passively sensitized with mouse monoclonal-IgE against AX, and humanized RBL-2H3 cells sensitized with polyclonal antibodies from sera of AX-allergic patients.
All BiAns were recognized by AX-sIgE. Dose-dependent activation responses were observed in both cellular assays, only with longer structures, containing spacers in the range of PEG 6000-12000 Da. Consistently, greater proportion of immunocomplexes and number of antibodies per complex for longerBiAns were visualized by TEM.
BiAns are valuable platforms to study the mechanism of effector cells activation. These nanomolecular toolshave demonstrated the importance of the adduct size to promote effector cells activation in AX allergy, which will impact for improvingin vitro diagnostics.

This article is protected by copyright. All rights reserved.

Author