Recognition and uptake of nanoparticle-conjugated food allergens by human monocytes

Stable biocompatible magnetic nanoparticles have demonstrated their potential in targeting biologically active molecules - including hormones, allergens, and various peptides - to specific cells or tissues expressing their cognate receptors, and may represent versatile and non-invasive tools for monitoring cellular functions both in vitro and in vivo. In this study, betalactoglobulin, one of the major whey proteins and food allergens, was covalently conjugated to biocompatible dextran-coated magnetic nanoparticles. The conjugated protein retained its structure and immunoreactivity towards monoclonal and polyclonal 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 adsorbed the allergen. To verify whether this approach could be complemented with others for improving our current understanding of the intracellular and intratissutal path of food allergens, BLG was conjugated to magnetic nanoparticles also labeled with a fluorescent probe. 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 direct evidence of actual internalization of the particles into the cytoplasm. These results open up the possibility to use a combination of these approaches for monitoring the fate of food allergens in cells that may be involved in the immune response to food

Biocompatible magnetic nanoparticles for protein cellular targeting

Various inorganic biocompatible nanoparticles (NPs) are considered suitable carriers for targeted cellular delivery of various drugs, proteins, and peptides through the cell membrane into cells. Stable NP conjugates may also be used to verify the uptake specificity and to monitor the post-uptake fate of the conjugated species. In this frame, covalent conjugates between carboxymethyl-dextran-coated magnetic NPs and bovine betalactoglobulin (BLG, a common food allergen) were prepared to address the issue of allergen recognition and uptake. NP biocompatibility was confirmed on various model cell systems. Then, human monocytes were incubated with different kinds of NPs, and subsequently separated by a magnetic system. BLG-functionalized NPs were taken up in larger amounts and more rapidly than unmodified NPs. Binding of fluorescent probes either to the NPs or to the NP-BLG conjugates allowed studies on their intracellular localization by confocal microscopy. These studies indicate that conjugates of common food allergens with magnetic NPs may be used to monitor the intracellular (and, ultimately, the intratissutal) path followed by food allergens when taken up by humans