Unravelling the lipocalin 2 interaction with aptamers: May rolling circle amplification improve their functional affinity?

Autor: Lorenzo-Gómez R; Dpto. Química Física y Analítica, Universidad de Oviedo, Julián Clavería 8, 33006 Oviedo, Spain; Instituto de Investigación Sanitaria del Principado de Asturias, Avenida de Roma, 33011 Oviedo, Spain., Fernández-Alonso N; Dpto. Química Física y Analítica, Universidad de Oviedo, Julián Clavería 8, 33006 Oviedo, Spain., Miranda-Castro R; Dpto. Química Física y Analítica, Universidad de Oviedo, Julián Clavería 8, 33006 Oviedo, Spain; Instituto de Investigación Sanitaria del Principado de Asturias, Avenida de Roma, 33011 Oviedo, Spain., de-Los-Santos-Álvarez N; Dpto. Química Física y Analítica, Universidad de Oviedo, Julián Clavería 8, 33006 Oviedo, Spain; Instituto de Investigación Sanitaria del Principado de Asturias, Avenida de Roma, 33011 Oviedo, Spain., Lobo-Castañón MJ; Dpto. Química Física y Analítica, Universidad de Oviedo, Julián Clavería 8, 33006 Oviedo, Spain; Instituto de Investigación Sanitaria del Principado de Asturias, Avenida de Roma, 33011 Oviedo, Spain. Electronic address: mjlc@uniovi.es.
Jazyk: angličtina
Zdroj: Talanta [Talanta] 2019 May 15; Vol. 197, pp. 406-412. Date of Electronic Publication: 2019 Jan 15.
DOI: 10.1016/j.talanta.2019.01.057
Abstrakt: Cancer diagnosis based on serum biomarkers requires receptors of extreme sensitivity and selectivity. Tunability of aptamer selection makes them ideal for that challenge. However, aptamer characterization is a time-consuming task, not always thoroughly addressed, leading to suboptimal aptamer performance. In this work, we report on the affinity characterization and potential usage of two aptamers against a candidate cancer biomarker, the neutrophil gelatinase-associated lipocalin (NGAL). Electrochemical sandwich assays on Au electrodes and SPR experiments showed a restricted capture ability of one of the aptamers (LCN2-4) and a small detectability of the other (LCN2-2). Interestingly, a truncated version of the signaling aptamer LCN2-2 selectively binds to NGAL covalently linked to magnetic beads due to high local protein concentration. The functional affinity of this aptamer is enhanced by three-orders of magnitude using rolling circle amplification (RCA), completed in only 15 min, followed by hybridization with short complementary fluorescein-tag probes, enzyme labeling and chronoamperometric measurement. Microscale thermophoresis experiments show a poor affinity for the protein in solution, which urges the importance of a full and in-depth characterization of aptamers to be used as diagnostic reagents.
(Copyright © 2019 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
Externí odkaz: