Droplet Surface Immunoassay by Relocation (D-SIRe) for High-Throughput Analysis of Cytosolic Proteins at the Single-Cell Level.
Fiche publication
Date publication
février 2023
Journal
Analytical chemistry
Auteurs
Membres identifiés du Cancéropôle Est :
Dr OULAD-ABDELGHANI Mustapha , Dr CHAUBET Guilhem
Tous les auteurs :
Dufossez R, Ursuegui S, Baudrey S, Pernod K, Mouftakhir S, Oulad-Abdelghani M, Mosser M, Chaubet G, Ryckelynck M, Wagner A
Lien Pubmed
Résumé
Enzyme-linked immunosorbent assay (ELISA) is a central analytic method in biological science for the detection of proteins. Introduction of droplet-based microfluidics allowed the development of miniaturized, less-consuming, and more sensitive ELISA assays by coencapsulating the biological sample and antibody-functionalized particles. We report herein an alternative in-droplet immunoassay format, which avoids the use of particles. It exploits the oil/aqueous-phase interface as a protein capture and detection surface. This is achieved using tailored perfluorinated surfactants bearing azide-functionalized PEG-based polar headgroups, which spontaneously react when meeting at the droplet formation site, with strained alkyne-functionalized antibodies solubilized in the water phase. The resulting antibody-functionalized inner surface can then be used to capture a target protein. This surface capture process leads to concomitant relocation at the surface of a labeled detection antibody and in turn to a drastic change in the shape of the fluorescence signal from a convex shape (not captured) to a characteristic concave shape (captured). This novel droplet surface immunoassay by fluorescence relocation (D-SIRe) proved to be fast and sensitive at 2.3 attomoles of analyte per droplet. It was further demonstrated to allow detection of cytosolic proteins at the single bacteria level.
Référence
Anal Chem. 2023 02 23;: