Silver nanoislands on cellulose fibers for chromatographic separation and ultrasensitive detection of small molecules

Abstract

High-throughput small-molecule assays play essential roles in biomedical diagnosis, drug discovery, environmental analysis, and physiological function research. Nanoplasmonics holds a great potential for the label-free detection of small molecules at extremely low concentrations. Here, we report the development of nanoplasmonic paper (NP-paper) for the rapid separation and ultrasensitive detection of mixed small molecules. NP-paper employs nanogap-rich silver nanoislands on cellulose fibers, which were simply fabricated at the wafer level by using low-temperature solid-state dewetting of a thin silver film. The nanoplasmonic detection allows for the scalable quantification and identification of small molecules over broad concentration ranges. Moreover, the combination of chromatographic separation and nanoplasmonic detection allows both the highly sensitive fluorescence detection of mixed small molecules at the attogram level and the label-free detection at the sub-nanogram level based on surface-enhanced Raman scattering. This novel material provides a new diagnostic platform for the high-throughput, low-cost, and label-free screening of mixed small molecules as an alternative to conventional paper chromatography.

Keywords: nanoplasmonics; paper chromatography; plasmon-enhanced spectroscopy; silver nanoislands; small-molecule assay.

Figure 1

Nanoplasmonic paper (NP-paper). (a) Large-area and batch nanofabrication of nanogap-rich silver nanoislands on cellulose fiber matrices using solid-state dewetting of a thin silver film. The thermally evaporated thin silver film on the cellulose fiber forms nanogap-rich silver nanoislands after thermal annealing at low temperatures. (b) A perspective-view scanning electron microscope (SEM) image of NP-paper (scale bar: 500 µm). The cellulose fibers possess a complex structural hierarchy from nanofibrils (20–200 nm in diameter) to microfibrils (5–20 µm in diameter). The inset shows a magnified SEM image of cellulose nano/microfibrils with abundant micropores (scale bar: 10 µm). (c) A cross-sectional SEM image of NP-paper. The top surface is only covered with nanogap-rich silver nanoislands, which maintain their hygroscopic nature and the physicochemical adsorption of the cellulose fiber matrices but strongly enhance light confinement on the surface (scale bar: 200 nm). (d) Photographic images of tailored normal chromatography paper (top), paper with thin silver film (middle), and ‘NP-paper’ with silver nanoislands (bottom) (scale bar: 10 mm).

Figure 2

Multiple EM hot spots from nanogap-rich silver nanoislands on cellulose fiber matrices. (a) Top-view SEM images (top) and electric field distributions derived from geometrical configurations of SEM images (bottom) of size-controlled silver nanoislands on cellulose fiber matrices (scale bar: 200 nm). The average diameter and the interstitial gap spacing of the silver nanoislands moderately increase with the initial silver film thickness (5 nm, 10 nm, 15 nm, and 20 nm) before thermal annealing. Based on the calculated FDTD results, the electric field at the LSPR wavelength (λ_LSPR) is locally enhanced by a factor of 24 for highly dense and nanogap-rich silver nanoislands formed from the initial Ag thickness of 10 nm. (b) The relationship between the surface mass coverages of the silver nanoislands and their extinction intensities measured at the individual plasmon resonance wavelengths. The maximum surface mass coverage and extinction at the LSPR wavelength are obtained from the initial silver thickness of 10 nm.

Figure 3

Ultrasensitive detection of small molecules. (a) Extinction at the absorption band (wavelength: 600 nm) of CV spotted on chromatography paper with (red solid rectangle) and without (black solid circle) silver nanoislands, depending on the solution concentration. For NP-paper, both the LSPR shift and the extinction quantitatively indicate the concentration of small molecules. CV molecules were detected from the extinction spectra for high concentrations ranging from 10 µM to 500 µM (optical micrographs: CV adsorbed on NP-paper) and even from the LSPR wavelength shift at lower concentrations from 50 nM to 10 µM (inset: λ_LSPR shift vs. the concentration), whereas normal paper can detect the extinction at over 10 µM in concentration. (b) Comparison of fluorescence emissions captured from FITC, SO, CR, and TB dye molecules adsorbed on paper without (top) and with (bottom) silver nanoislands (scale bar: 200 µm). (c) Fluorescence enhancement of FITC, SO, CR, and TB molecules at 500 nM. The fluorescence intensities are increased by a factor of 8 for TB, and the enhancement ratios for the individual molecules strongly correlate with the reflectance spectra of the silver nanoislands measured from NP-paper.

Figure 4

Colorimetric, MEF, and SERS detections after small-molecule separation. (a) Colorimetric detection of dye molecules (CR, TB, and CV) in an aqueous mixture after chromatographic separation on NP-paper. The separated molecules of CR, TB, and CV at the 1, 2, and 3 positions, respectively, are clearly distinguished and also confirmed by the corresponding absorption peaks. The SERS measurement also enables label-free detection after chromatographic separation. (b) Fluorescence detection of mixed dye molecules at ultra-low concentration after the chromatographic separation. The fluorescence signals scanned along the column successfully demonstrate the clear partitioning of SO, TB, and CR dye molecules at sub-picogram levels adsorbed on NP-paper. The retention factors of CR, TB, and SO are 0.13, 0.48, and 0.96, respectively. (c) Chromatographic SERS of mixed vitamins (one water-soluble riboflavin of 10 ng and two fat-soluble β-carotene of 537 pg and α-tocopherol of 11 ng). The SERS spectra of the vitamins are plotted at their maxima on the intensity map. The inset indicates the fluorescence micrographs of riboflavin measured at 530–600 nm in wavelength (scale bar: 300 µm).