Detection of miRNA cancer biomarkers using light activated Molecular Beacons.

Autor:	Gonçalves OSL; Medical Photonics Lab, Department of Health Science and Technology, Faculty of Medicine, Aalborg University Fredrik Bajers Vej 7 DK-9220 Aalborg Denmark., Wheeler G; School of Biological Sciences, University of East Anglia Norwich Research Park Norwich NR4 7TJ UK., Dalmay T; School of Biological Sciences, University of East Anglia Norwich Research Park Norwich NR4 7TJ UK., Dai H; Biosynthesis 612 East Main Street Lewisville TX 75057-4052 USA., Castro M; Biosynthesis 612 East Main Street Lewisville TX 75057-4052 USA., Castro P; Biosynthesis 612 East Main Street Lewisville TX 75057-4052 USA., García-Rupérez J; Nanophotonics Technology Center, Universitat Politècnica de València Camino de Vera S/N 46022 Valencia Spain., Ruiz-Tórtola Á; Nanophotonics Technology Center, Universitat Politècnica de València Camino de Vera S/N 46022 Valencia Spain., Griol A; Nanophotonics Technology Center, Universitat Politècnica de València Camino de Vera S/N 46022 Valencia Spain., Hurtado J; Nanophotonics Technology Center, Universitat Politècnica de València Camino de Vera S/N 46022 Valencia Spain., Bellieres L; Nanophotonics Technology Center, Universitat Politècnica de València Camino de Vera S/N 46022 Valencia Spain., Bañuls MJ; IDM, Instituto Interuniversitario de Investigación de Reconocimiento Molecular y Desarrollo Tecnológico, Departamento de Química, Universitat Politècnica de València Camino de Vera S/N 46022 Valencia Spain., González D; IDM, Instituto Interuniversitario de Investigación de Reconocimiento Molecular y Desarrollo Tecnológico, Departamento de Química, Universitat Politècnica de València Camino de Vera S/N 46022 Valencia Spain., López-Guerrero JA; Biología Molecular, Fundación Instituto Valenciano de Oncología Valencia Spain., Neves-Petersen MT; Department of Clinical Medicine, Aalborg University Hospital Hobrovej 18-22 9000 Aalborg Denmark nevespetersen@gmail.com +45 2252 2475.; Department of Biomedical Sciences and Medicine, Centre for Biomedical Research (CBMR), University of Algarve, Campus de Gambelas 8005-139 Faro Portugal.
Jazyk:	angličtina
Zdroj:	RSC advances [RSC Adv] 2019 Apr 25; Vol. 9 (22), pp. 12766-12783. Date of Electronic Publication: 2019 Apr 25 (Print Publication: 2019).
DOI:	10.1039/c9ra00081j
Abstrakt:	Early detection of cancer biomarkers can reduce cancer mortality rate. miRNAs are small non-coding RNAs whose expression changes upon the onset of various types of cancer. Biosensors that specifically detect such biomarkers can be engineered and integrated into point-of-care devices (POC) using label-free detection, high sensibility and compactness. In this paper, a new engineered Molecular Beacon (MB) construct used to detect miRNAs is presented. Such a construct is immobilized onto biosensor surfaces in a covalent and spatially oriented way using the photonic technology Light Assisted Molecular Immobilization (LAMI). The construct consists of a Cy3 labelled MB covalently attached to a light-switchable peptide. One MB construct contains a poly-A sequence in its loop region while the other contains a sequence complementary to the cancer biomarker miRNA-21. The constructs have been characterized by UV-Vis spectroscopy, mass spectrometry and HPLC. LAMI led to the successful immobilization of the engineered constructs onto thiol functionalized optically flat quartz slides and Silicon on Insulator (SOI) sensor surfaces. The immobilized Cy3 labelled MB construct has been imaged using confocal fluorescence microscopy (CFM). The bioavailability of the immobilized engineered MB biosensors was confirmed through specific hybridization with the Cy5 labelled complementary sequence and imaged by CFM and FRET. Hybridization kinetics have been monitored using steady state fluorescence spectroscopy. The label-free detection of miRNA-21 was also achieved by using integrated photonic sensing structures. The engineered light sensitive constructs can be immobilized onto thiol reactive surfaces and are currently being integrated in a POC device for the detection of cancer biomarkers. Competing Interests: The authors declare no conflicts of interest. (This journal is © The Royal Society of Chemistry.)
Databáze:	MEDLINE
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