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Direct Quantification of MicroRNA at Low Picomolar Level in Sera of Glioma Patients Using a Competitive Hybridization Followed by Amplified Voltammetric Detection

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Download Hi-Res ImageDownload to MS-PowerPointCite This:Anal. Chem. 2012, 84, 15, 6400-6406
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    Direct Quantification of MicroRNA at Low Picomolar Level in Sera of Glioma Patients Using a Competitive Hybridization Followed by Amplified Voltammetric Detection
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    † College of Chemistry and Chemical Engineering, Central South University, Changsha, Hunan, People’s Republic of China 410083
    ‡ Cancer Research Institute, Central South University, Changsha, Hunan, People’s Republic of China 410013
    § Department of Chemistry and Biochemistry, California State University, Los Angeles, Los Angeles, California 90032, United States
    *E-mail: [email protected] (J.W.); [email protected] (M.W.); [email protected] (F.Z.).
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    Analytical Chemistry

    Cite this: Anal. Chem. 2012, 84, 15, 6400–6406
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    https://doi.org/10.1021/ac203368h
    Published June 26, 2012
    Copyright © 2012 American Chemical Society
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    MicroRNAs (miRNAs), acting as oncogenes or tumor suppressors in humans, play a key role in regulating gene expression and are believed to be important for developing novel therapeutic treatments and clinical prognoses. Due to their short lengths (17–25 nucleotides) and extremely low concentrations (typically < picomolar) in biological samples, quantification of miRNAs has been challenging to conventional biochemical methods, such as Northern blotting, microarray, and quantitative polymerase chain reaction (qPCR). In this work, a biotinylated miRNA (biotin-miRNA) whose sequence is the same as that of a miRNA target is introduced into samples of interest and allowed to compete with the miRNA target for the oligonucleotide (ODN) probe preimmobilized onto an electrode. Voltammetric quantification of the miRNA target was accomplished after complexation of the biotin-miRNA with ferrocene (Fc)-capped gold nanoparticle/streptavidin conjugates. The Fc oxidation current was found to be inversely proportional to the concentration of target miRNA between 10 fM and 2.0 pM. The method is highly reproducible (relative standard deviation (RSD) < 5%), regenerable (at least 8 regeneration/assay cycles without discernible signal decrease), and selective (with sequence specificity down to a single nucleotide mismatch). The low detection levels (10 fM or 0.1 attomoles of miRNA in a 10 μL solution) allow the direct quantification of miRNA-182, a marker correlated to the progression of glioma in patients, to be performed in serum samples without sample pretreatment and RNA extraction and enrichment. The concentration of miRNA-182 in glioma patients was found to be 3.1 times as high as that in healthy persons, a conclusion in excellent agreement with a separate qPCR measurement of the expression level. The obviations of the requirement of an internal reference in qPCR, simplicity, and cost-effectiveness are other additional advantages of this method for detection of nucleic acids in clinical samples.

    Copyright © 2012 American Chemical Society

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    This article is cited by 98 publications.

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    Cite this: Anal. Chem. 2012, 84, 15, 6400–6406
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    https://doi.org/10.1021/ac203368h
    Published June 26, 2012
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