High throughput single nanoparticle spectroscopy

Abstract

Progress in the development and application of nanoengineered systems is limited by the availability of quantitative measurement techniques. For the engineering of nanoparticle (NP)-based systems, single NP characterization is essential, but existing methods are slow and low throughput. We demonstrate a flow spectroscopy technique capable of analyzing hundreds of nanoparticles per second and use this technique for the high throughput analysis of nanoparticle surface-enhanced resonant Raman scattering (SERRS) tags. By measuring Rayleigh and Raman scattering from thousands of individual tags, tag preparations can be characterized based on their brightness and uniformity. The rapid analysis of individual nanoparticles using high spectral resolution flow spectroscopy will be useful in many areas of nanoengineering.

Figure 1

Single nanoparticle flow spectrometer. (a) Sample is hydrodynamically focused to flow through a focused laser beam. Note that the figure is not to scale. (b) Schematic of the Raman flow spectrometer setup. Refer to the Materials and Methods section for detailed instrument information. (c) Single tag Raman scattering spectra of an inactive SERRS tag (black) and Raman-active tags prepared with oxazine 170 (red), thionin (blue), and malachite green (green) of varying intensities. Spectra have been offset for clarity.

Figure 2

Ensemble and flow spectroscopy analysis of nanoshell SERRS tags. (a) Ensemble oxazine 170 SERRS tag Raman spectrum from tags prepared with 2 μL of silver enhancement solution. Spectrum was collected from a 0.9 pM nanoshell solution (5.5 × 10⁸ NPs/mL as measured using flow spectroscopy) using 14 mW of 647 nm laser excitation and a signal integration time of 2 s. (b) Univariate and bivariate displays of Rayleigh and Raman scattering intensity from single tag flow spectroscopy analysis and representative single tag spectra from each subpopulation. Spectra were collected using 340 mW of 647 nm excitation and signal integration times of 300 μs and have been offset for clarity.

Figure 3

SERRS tag population analysis. (a) Histograms of Raman scattering intensity from three SERRS tag preparations prepared with 0, 3, and 5 μL of silver enhancement solution. Illustrations display the relative sizes of the silica core (gray), gold shell (yellow), and silver shell (blue). (b–d) Density plots of SERRS tags prepared with 0, 3, and 5 μL of silver enhancement solution. The analysis was performed using 340 mW of 647 nm excitation and signal integration times of 300 μs.

Figure 4

Analysis of SERRS tag preparations. (a) Mean Raman scattering intensity of Raman-active tags (red squares) for each tag preparation and the percentage of Raman-active tags (black circles) were found to vary with the thickness of the tag silver layer. (b) Representative TEM images of SERRS tags with silver layers of varying thickness. Scale bar is 500 nm.