[en] Lyme borreliosis (LB) is the most prevalent tick-borne illness, with an estimated 700 000 cases annually in the United States and Europe. The LB diagnosis based on a two-tiered serology remains controversial due to its indirect nature and low sensitivity during the early stage of the disease. Aptamers are single-stranded DNA or RNA oligonucleotides that exhibit high selectivity and specificity for their target due to their unique three-dimensional structure. By applying cross-over-SELEX process, an enrichment of DNA oligonucleotide sequences against a surface protein of Borrelia, named CspZ, has been performed and monitored using absorbance at 260 nm, melting curves and NGS analyses. Beyond sequence enrichment, oligonucleotides binding to CspZ were observed during the selection rounds by Dot Blot and beads assays. Thirteen unique and highly redundant oligonucleotide sequences were further characterized using multiple approaches such as Dot Blot, BioLayer Interferometry and Surface Plasmon Resonance. The selected aptamers showed KD values from tens of nanomolar to the micromolar range by BLI and SPR. Two aptamers, Apta9 and Apta10, characterized by flow cytometry and epifluorescence microscopy, were able to specifically recognize Borrelia burgdorferi sensu stricto. This strategy holds promise for the development of an improved diagnostic assay.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Guérin, Mickaël; Unité de Génie Enzymatique et Cellulaire (GEC), CNRS UMR 7025, Université de Technologie de Compiègne, 60203, Compiègne, France
Matagne, André ; Université de Liège - ULiège > Département des sciences de la vie > Enzymologie et repliement des protéines
Aucher, Willy; Laboratoire Ecologie & Biologie des Interactions (EBI), CNRS UMR 7267, Université de Poitiers, 86073, Poitiers, France
Verdon, Julien; Laboratoire Ecologie & Biologie des Interactions (EBI), CNRS UMR 7267, Université de Poitiers, 86073, Poitiers, France
Paoli, Emmeline; Laboratoire Ecologie & Biologie des Interactions (EBI), CNRS UMR 7267, Université de Poitiers, 86073, Poitiers, France
Ducrotoy, Jules; Unité de Génie Enzymatique et Cellulaire (GEC), CNRS UMR 7025, Université de Technologie de Compiègne, 60203, Compiègne, France
Octave, Stéphane ; Unité de Génie Enzymatique et Cellulaire (GEC), CNRS UMR 7025, Université de Technologie de Compiègne, 60203, Compiègne, France
Avalle, Bérangère; Unité de Génie Enzymatique et Cellulaire (GEC), CNRS UMR 7025, Université de Technologie de Compiègne, 60203, Compiègne, France
Maffucci, Irene; Unité de Génie Enzymatique et Cellulaire (GEC), CNRS UMR 7025, Université de Technologie de Compiègne, 60203, Compiègne, France
Padiolleau-Lefèvre, Séverine ; Unité de Génie Enzymatique et Cellulaire (GEC), CNRS UMR 7025, Université de Technologie de Compiègne, 60203, Compiègne, France. severine.padiolleau@utc.fr
Language :
English
Title :
Selection and characterization of DNA aptamers targeting the surface Borrelia protein CspZ with high-throughput cross-over SELEX.
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