Uptake of nanoparticle-conjugated allergens by human monocytes

INTRODUCTION: Stable biocompatible magnetic nanoparticles, conveniently grafted with affinity ligands and/or transfection agents to integrate functionally and efficiently into cellular and subcellular structures, can confer cells magnetic activity, and may represent versatile and non-invasive tools for targeting specific cellular structures, and for monitoring cellular functions both in vitro and in vivo. RESULTS: In this study, betalactoglobulin, the most abundant protein in bovine whey and a major food allergens, was covalently conjugated to biocompatible carboxymethyldextran-coated magnetic nanoparticles. The conjugated protein retained its immunoreactivity towards different monoclonal antibodies. BLG-conjugated nanoparticles were taken up by human monocytes much more efficiently than non-conjugated particles, allowing easy magnetic separation of cells that had integrated the allergen. To assess whether the interaction between monocytes and the nanoparticles was associated to their internalization rather than to mere adsorption of the particles on the cell surface, BLG was conjugated to fluorescent-labeled magnetic nanoparticles. The uptake of these materials by human monocytes was monitored through flow cytometry and confocal microscopy, in comparison with fluorescent nanoparticles devoid of the allergen or conjugated with human serum albumin. Both approaches confirm a higher uptake of the BLG-conjugated particles, and confocal microscopy provided clear evidence of particles internalization into the cytoplasm. These results pave the way to use a combination of these approaches to improve the current understanding of the intracellular and intratissutal path of allergens that may be involved in the immune response to food.