Novel fluorescence assay for tracking molecular and cellular allergen-protein interactions.

T cells recognizing nickel (Ni) are key mediators in human Ni allergy, which represents the most common form of human contact hypersensitivity. In contrast to well-characterized Ni-specific human T cell clones, molecular knowledge about the extra- and intracellular route(s) of antigen/allergen presentation and processing of Ni-specific epitopes is still fragmentary. Here, we demonstrate a new metal-specific fluorescent technique to detect and quantify metal ions, like Ni(2+), while they are associated with isolated metalloproteins. Moreover, utilizing the fluorescent metal sensor molecule Newport Green (NPG) a novel method has been developed, which permits the metal-specific detection of Ni(2+) binding to surface or intracellular structures of individual human antigen presenting cells by flow cytometry. We expect such metal-specific fluorescent analyses to contribute to a better basic understanding of molecular and cellular immune processes involved in Ni-specific T cell epitope generation and the pathogenesis of human nickel allergy.