In recent years, biomarkers that can be used for minimally invasive diagnosis “liquid biopsy” have attracted attention. Circulating tumor DNA (ctDNA), circulating tumor RNA (ctRNA) and circulating tumor cells (CTC) in body fluids are used as biomarkers for liquid biopsy because expression levels of them in body fluids are associated with the onset of the diseases. Although quantitative PCR is generally used to quantify nucleic acids, the PCR has been poorly utilized as a diagnostic method due to complicated operations for amplification of nucleic acids. Therefore, a nucleic acid detection system that does not require the amplification steps of nucleic acids has been desired. Surface-enhanced Raman scattering (SERS) has gained momentum as an important tool in analytical chemistry since its discovery in 1974. The Raman signal derived from the molecule adsorbed on the metal surface is enhanced from 108 to 1010 times by SERS. Until now, we have developed a SERS probe containing a gold nanorod as a SERS substrate and chemical tags with a Raman signal in a region where the Raman signals from cellular components are negligible and constructed a sandwich-typed assay for the nucleic acids using the SERS probes. First, we prepared gold nanorods with alkylthiol-capped 20-mer oligonucleotides and chemical tags (SERS probes). On average, ca. 240 oligonucleotides are introduced on each gold nanorod. Raman spectra of the SERS probes were directly measured using a Raman microscope. As a result of Raman measurement, a Raman signal derived from the chemical tag was observed in the silent region. Next, sandwich-typed assays for detection of a target DNA were performed using the SERS probes and a significant Raman peak was observed in the presence of the target DNA, whereas no Raman signals from the tag were observed in the absence of the target DNA. Furthermore, to evaluate the selectivity of the system, sandwich-typed assays were conducted using two kinds of target DNAs. From the Raman spectra, two kinds of target DNAs were discriminated by the sandwich-typed assay.

Development of a sandwich-typed assay for nucleic acids using surface-enhanced Raman scattering

256th National Meeting (2018)

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