Luminescent lanthanide nanoparticles for ultrasensitive Enzyme-Linked Immunosorbent Assay 'ELISA'

In this thesis, we developed a new ELISA method based on luminescent Ln-NPs to overcome the limitations of commercial detection methods based on organic chromophores or fluorophores. The principle was to employ an enzymatic reaction to convert a substrate which cannot photosensitize the NPs into an active form that can be coordinated to the surface and efficiently sensitize Ln³⁺ ions. A first part focused on the synthesis and the characterization of Tb and Tb/Eu NPs. Then, two approaches were adopted to develop the enzymatic reaction. The first consisted in the hydroxylation of isophthalate derivatives catalyzed by HRP to form the 2-hydroxy-isophthalate, an efficient sensitization unit of the NPs. The second consisted in the ortho-dimerization of phenols catalyzed by the same enzyme, while the substrate was optimized to promote the formation of an efficient NPs sensitizer. The latter provided very promising results in terms of sensitivity of HRP detection in solution. The developed system was then implanted in a full ELISA for insulin detection, where we demonstrated a sensitivity improvement compared to the colorimetric detection method of the commercial kit. (author)