title: Application of Spectral Crosstalk Correction for Improving Multiplexed MicroRNA Detection Using a Single Excitation Wavelength.
abstract: MicroRNAs (miRNAs) play crucial roles in the regulation of cellular activities and are next-generation biomarkers for early cancer detection. Simultaneous monitoring of multiplexed miRNA is very important for enhancing the accuracy of cancer diagnostics. Traditional fluorescence methods for multicomponent analysis were usually operated under multiple excitation wavelengths, because spectral crosstalk is very detrimental to detecting accuracy for multicomponent analysis. Herein, we present a fluorescence strategy for multi-miRNAs detection in plasma under a single excitation wavelength. Nucleic acid stain TOTO-1 and three labeled fluorescence dyes Cy3, Cy3.5, and Cy5 emit no fluorescence in their free state. Target miRNA hybridized the auxiliary and probe oligonucleotides into duplex nucleic acid. Intercalation interaction localized TOTO-1 and labeled dyes into the duplex nucleic acid. As a result, TOTO-1 emitted strong fluorescence and efficient Förster resonance energy transfer (FRET) happened. MicroRNAs miRNA-155, miRNA-182, and miRNA-197, which are significant for the early diagnosis of lung cancer, were simultaneously detected as models. Deviations from spectral crosstalk in the presence of other miRNAs were corrected by mathematical methods. Results demonstrated that, after spectra crosstalk corrections, every miRNA at high or low concentration in plasma was determined accurately in the presence of either high or low concentrations of the other two miRNAs. This new multiplexed assay for miRNAs is promising for clinical diagnosis, prognosis, and therapeutic monitoring of early-stage lung cancer.
authors.affiliation: Jiangsu Engineering Laboratory of Smart Carbon-Rich Materials and Device, Laboratory of Environmental Medicine Engineering, Ministry of Education, School of Chemistry and Chemical Engineering, Southeast University , Nanjing, 211189, China.
authors.forename: Yuanjian
authors.initials: Y
authors.lastname: Liu
authors.affiliation: College of Food Science and Technology, Henan University of Technology , Zhengzhou, 450001, China.
authors.forename: Min
authors.initials: M
authors.lastname: Wei
authors.affiliation: Jiangsu Engineering Laboratory of Smart Carbon-Rich Materials and Device, Laboratory of Environmental Medicine Engineering, Ministry of Education, School of Chemistry and Chemical Engineering, Southeast University , Nanjing, 211189, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Li
authors.affiliation: Jiangsu Engineering Laboratory of Smart Carbon-Rich Materials and Device, Laboratory of Environmental Medicine Engineering, Ministry of Education, School of Chemistry and Chemical Engineering, Southeast University , Nanjing, 211189, China.
authors.forename: Anran
authors.initials: A
authors.lastname: Liu
authors.affiliation: Jiangsu Engineering Laboratory of Smart Carbon-Rich Materials and Device, Laboratory of Environmental Medicine Engineering, Ministry of Education, School of Chemistry and Chemical Engineering, Southeast University , Nanjing, 211189, China.
authors.forename: Wei
authors.identifier: 0000-0003-1893-1105
authors.initials: W
authors.lastname: Wei
authors.affiliation: Jiangsu Engineering Laboratory of Smart Carbon-Rich Materials and Device, Laboratory of Environmental Medicine Engineering, Ministry of Education, School of Chemistry and Chemical Engineering, Southeast University , Nanjing, 211189, China.
authors.forename: Yuanjian
authors.identifier: 0000-0003-2932-4159
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Jiangsu Engineering Laboratory of Smart Carbon-Rich Materials and Device, Laboratory of Environmental Medicine Engineering, Ministry of Education, School of Chemistry and Chemical Engineering, Southeast University , Nanjing, 211189, China.
authors.forename: Songqin
authors.identifier: 0000-0002-4686-5291
authors.initials: S
authors.lastname: Liu
mesh_terms: D004247:DNA / Q000737:chemistry; D055088:Early Detection of Cancer; D005453:Fluorescence; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry*; D006801:Humans; D008175:Lung Neoplasms / Q000097:blood / Q000000981:diagnostic imaging*; D035683:MicroRNAs / Q000097:blood*; D013844:Thiazoles / Q000737:chemistry*
chemical_list: D005456:Fluorescent Dyes; D035683:MicroRNAs; C086329:TOTO-1 thiazole orange dye; D013844:Thiazoles; D004247:DNA