Despite the need in various applications, accurate quantification of nucleic acids still remains a challenge. The widely-used qPCR has reduced accuracy at ultralow template concentration and is susceptible to nonspecific amplifications. The more recently developed dPCR is costly and cannot handle high-concentration samples. We combine the strengths of qPCR and dPCR by performing PCR in silicon-based microfluidic chips and demonstrate high quantification accuracy in a large concentration range. Importantly, at low template concentration, we observe on-site PCR (osPCR), where only certain sites of the channel show amplification. The sites have almost identical ct values, showing osPCR is a quasi-single molecule phenomenon. Using osPCR, we can measure both the ct values and the absolute concentration of templates in the same reaction. Additionally, osPCR enables identification of each template molecule, allowing removal of nonspecific amplification during quantification and greatly improving quantification accuracy. We develop sectioning algorithm that improves the signal amplitude and demonstrate improved detection of COVID in patient samples.

Surface-enhanced Raman scattering (SERS) as a high sensitivity analytical method for molecule detection has attracted much attention in recent research. In this work, we demonstrated an improved SERS substrate, which has the gold nanoparticles randomly distributed on a SiO₂ interception layer over a gold thin film layer on the flat sapphire substrate (AuNP/SiO₂/Au/Sapphire), over the dispersed gold nanoparticles on a silicon substrate (AuNP/Si), for detection of R6G (1 × 10⁻⁶ M) in a Raman microscope. The fabrication of sandwich layers on top of the sapphire substrate involves evaporation of a gold mirror as thick as 100 nm, plasma enhanced chemical vapor deposition of the silica insulator layer 10 nm thick, and evaporation of a thin gold layer 10 nm thick for forming gold nanoparticles. For comparison, a gold thin film with a thickness of 5 nm and 10 nm was evaporated on a silicon substrate, respectively (AuNP/Si), as the reference SERS substrates in the experiment. The AuNP/SiO₂/Au/Sapphire substrate demonstrated improved sensitivity in detection of molecules in Raman microscopy, which can enable the molecules to be recognizable at a low laser power as 8.5 × 10⁻³ mW, 0.017 mW, 0.085 mW, and 0.17 mW for ultrashort exposure time. The simulation of AuNP/SiO₂/Au/Sapphire substrate and AuNP/Si substrate, based on the finite-difference time-domain (FDTD) method, explained the improved sensitivity for detection of R6G molecules from the view of classical electromagnetics, and it suggested the optimized size for the gold nanoparticles and the optimized laser wavelength for Raman microscopy for further research.

Plasmonic metallic nanostructures have attracted much interest for their ability to manipulate light on a subwavelength scale and for their related applications in various fields. In this work, a parabola-like gold nanobowtie (PGNB) on a sapphire substrate was designed as a nano-cavity for confining light waves in a nanoscale gap region. The near-field optical properties of the innovative PGNB structure were studied comprehensively, taking advantage of the time-resolved field calculation based on a finite-difference time-domain algorithm (FDTD). The calculation result showed that the resonance wavelength of the nano-cavity was quite sensitive to the geometry of the PGNB. The values that related to the scattering and absorption properties of the PGNB, such as the scattering cross section, absorption cross section, extinction cross section, scattering ratio, and also the absorption ratio, were strongly dependent on the geometrical parameters which affected the surface area of the nanobowtie. Increased sharpness of the gold tips on the parabola-like nano-wings benefited the concentration of high-density charges with opposite electric properties in the narrow gold tips with limited volume, thus, resulting in a highly enhanced electric field in the nano-cavity under illumination of the light wave. Reduction of the gap size between the two gold nano-tips, namely, the size of the nano-cavity, decreased the distance that the electric potential produced by the highly concentrated charges on the surface of each gold nano-tip had to jump across, therefore, causing a significantly enhanced field in the nano-cavity. Further, alignment of the linearly polarized electric field of the incident light wave with the symmetric axis of the PGNB efficiently enabled the free electrons in the PGNB to concentrate on the surface of the sharp gold tips with a high density, thus, strongly improving the field across the nano-cavity. The research provides a new insight for future design, nanofabrication, and characterization of PGNBs for applications in devices that relate to enhancing photons emission, improving efficiency for energy harvesting, and improving sensitivity for infrared detection.