title: ELAKCA: Enzyme-Linked Aptamer Kissing Complex Assay as a Small Molecule Sensing Platform.
abstract: We report herein a novel sandwich-type enzyme-linked assay for the "signal-on" colorimetric detection of small molecules. The approach (referred to as enzyme-linked aptamer kissing complex assay (ELAKCA)) relied on the kissing complex-based recognition of the target-bound hairpin aptamer conformational state by a specific RNA hairpin probe. The aptamer was covalently immobilized on a microplate well surface to act as target capture element. Upon small analyte addition, the folded aptamer was able to bind to the biotinylated RNA hairpin module through loop-loop interaction. The formed ternary complex was then revealed by the introduction of the streptavidin-horseradish peroxidase conjugate that catalytically converted the 3,3',5,5'-tetramethylbenzidine substrate into a colorimetric product. ELAKCA was successfully designed for two different systems allowing detecting the adenosine and theophylline molecules. The potential practical applicability in terms of biological sample analysis (human plasma), temporal stability, and reusability was also reported. Owing to the variety of both hairpin functional nucleic acids, kissing motifs, and enzyme-based signaling systems, ELAKCA opens up new prospects for developing small molecule sensing platforms of wide applications.
authors.affiliation: University Grenoble Alpes, DPM UMR 5063, F-38041 Grenoble, CNRS, DPM UMR 5063, F-38041 Grenoble, France.|Département de Biochimie, Toxicologie et Pharmacologie, CHU de Grenoble Site Nord - Institut de Biologie et de Pathologie , F-38041 Grenoble, France.
authors.forename: Benoit
authors.initials: B
authors.lastname: Chovelon
authors.affiliation: Laboratoire ARNA, Université Bordeaux, Inserm U869, F-33076 Bordeaux, France.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Durand
authors.affiliation: Laboratoire ARNA, Université Bordeaux, Inserm U869, F-33076 Bordeaux, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Dausse
authors.affiliation: Laboratoire ARNA, Université Bordeaux, Inserm U869, F-33076 Bordeaux, France.
authors.forename: Jean-Jacques
authors.initials: JJ
authors.lastname: Toulmé
authors.affiliation: Département de Biochimie, Toxicologie et Pharmacologie, CHU de Grenoble Site Nord - Institut de Biologie et de Pathologie , F-38041 Grenoble, France.|University Grenoble Alpes, Laboratory of Hypoxy Physiopathology Study Inserm U1042, 38700 La Tronche, France.
authors.forename: Patrice
authors.initials: P
authors.lastname: Faure
authors.affiliation: University Grenoble Alpes, DPM UMR 5063, F-38041 Grenoble, CNRS, DPM UMR 5063, F-38041 Grenoble, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Peyrin
authors.affiliation: University Grenoble Alpes, DPM UMR 5063, F-38041 Grenoble, CNRS, DPM UMR 5063, F-38041 Grenoble, France.
authors.forename: Corinne
authors.initials: C
authors.lastname: Ravelet
mesh_terms: D000241:Adenosine / Q000097:blood; D052157:Aptamers, Nucleotide / Q000737:chemistry; D001560:Benzidines / Q000737:chemistry; D002863:Chromogenic Compounds / Q000737:chemistry; D003124:Colorimetry; D057075:Enzyme Assays / Q000295:instrumentation / Q000379:methods*; D006735:Horseradish Peroxidase / Q000737:chemistry; D006801:Humans; D019809:Streptavidin / Q000737:chemistry; D013806:Theophylline / Q000097:blood
chemical_list: D052157:Aptamers, Nucleotide; D001560:Benzidines; D002863:Chromogenic Compounds; C021758:3,3',5,5'-tetramethylbenzidine; D019809:Streptavidin; D013806:Theophylline; D006735:Horseradish Peroxidase; D000241:Adenosine