Tip-Enhanced Raman Imaging of Single-Stranded DNA with Single Base Resolution

Abstract

Tip-enhanced Raman scattering (TERS) is a promising optical and analytical technique for chemical imaging and sensing at single molecule resolution. In particular, TERS signals generated by a gap-mode configuration where a silver tip is coupled with a gold substrate can resolve a single-stranded DNA (ssDNA) molecule with a spatial resolution below 1 nm. To demonstrate the proof of subnanometer resolution, we show direct nucleic acid sequencing using TERS of a phage ssDNA (M13mp18). M13mp18 provides a known sequence and, through our deposition strategy, can be stretched (uncoiled) and attached to the substrate by its phosphate groups, while exposing its nucleobases to the tip. After deposition, we scan the silver tip along the ssDNA and collect TERS signals with a step of 0.5 nm, comparable to the bond length between two adjacent DNA bases. By demonstrating the real-time profiling of a ssDNA configuration and furthermore, with unique TERS signals of monomeric units of other biopolymers, we anticipate that this technique can be extended to the high-resolution imaging of various nanostructures as well as the direct sequencing of other important biopolymers including RNA, polysaccharides, and polypeptides.

Fabrication of a Biocompatible Mica/Gold Surface for Tip‐Enhanced Raman Spectroscopy

“…This application of TERS has thus far been limited, due to difficulties in generating high field enhancements while maintaining biocompatibility. [4][5][6][7][8][9][10][11][12] However, there remain significant challenges to its widespread implementation and capacity to reliably address biological and clinical questions.The sensitivity of TERS arises from the excitation of a localized plasmon resonance in a noble metal AFM tip, which can provide an order of magnitude enhancement in the optical field close to the tip. However, metals have poor biocompatibility, potentially introducing difficulties in characterizing native structure and conformation in biomolecules, whereas biocompatible surfaces have weak optical field enhancements.…”

“…The high sensitivity achievable through TERS arises from the excitation of a localized surface plasmon resonance in a noble metal atomic force microscope (AFM) tip, which in combination with a metallic surface can produce huge enhancements in the local optical field. Most TERS studies are additionally performed on noble metal surfaces, [8,[12][13][14] where plasmonic coupling between tip and surface further increases the optical field enhancement. Herein, a novel, biocompatible, highly enhancing surface is designed and fabricated based on few-monolayer mica flakes, mechanically exfoliated on a metal surface.…”

“…These surfaces allow the formation of coupled plasmon enhancements for TERS imaging, while maintaining the biocompatibility and atomic flatness of the mica surface for high resolution AFM. [12,19] For example, surface-enhanced Raman spectroscopy (SERS) studies have revealed that the interactions with the plasmonic metal surface promote protein denaturation and DNA double strand breaks. We demonstrate up to five orders of magnitude improvement in TERS signals over conventional mica surfaces, expanding the sensitivity of TERS to a wide range of non-resonant biomolecules with weak Raman cross-sections.…”

“…AFM is a powerful technique for imaging on the single-molecule level, [1] and Raman spectroscopy can distinguish biological constituents and their organization and conformation through their vibrational signatures. [4][5][6][7][8][9][10][11][12] However, there remain significant challenges to its widespread implementation and capacity to reliably address biological and clinical questions. [4][5][6][7][8][9][10][11][12] However, there remain significant challenges to its widespread implementation and capacity to reliably address biological and clinical questions.…”

“…Most TERS studies are additionally performed on noble metal surfaces, [8,[12][13][14] where plasmonic coupling between tip and surface further increases the optical field enhancement. Most TERS studies are additionally performed on noble metal surfaces, [8,[12][13][14] where plasmonic coupling between tip and surface further increases the optical field enhancement.…”