Unique temporal and spatial biomolecular emission profile on individual zinc oxide nanorods

Abstract

Zinc oxide nanorods (ZnO NRs) have emerged in recent years as extremely useful, optical signal-enhancing platforms in DNA and protein detection. Although the use of ZnO NRs in biodetection has been demonstrated so far in systems involving many ZnO NRs per detection element, their future applications will likely take place in a miniaturized setting while exploiting single ZnO NRs in a low-volume, high-throughput bioanalysis. In this paper, we investigate temporal and spatial characteristics of the biomolecular fluorescence on individual ZnO NR systems. Quantitative and qualitative examinations of the biomolecular intensity and photostability are carried out as a function of two important criteria, the time and position along the long axis (length) of NRs. Photostability profiles are also measured with respect to the position on NRs and compared to those characteristics of biomolecules on polymeric control platforms. Unlike the uniformly distributed signal observed on the control platforms, both the fluorescence intensity and photostability are position-dependent on individual ZnO NRs. We have identified a unique phenomenon of highly localized, fluorescence intensification on the nanorod ends (FINE) of well-characterized, individual ZnO nanostructures. When compared to the polymeric controls, the biomolecular fluorescence intensity and photostability are determined to be higher on individual ZnO NRs regardless of the position on NRs. We have also carried out finite-difference time-domain simulations the results of which are in good agreement with the observed FINE. The outcomes of our investigation will offer a much needed basis for signal interpretation for biodetection devices and platforms consisting of single ZnO NRs and, at the same time, contribute significantly to provide insight in understanding the biomolecular fluorescence observed from ZnO NR ensemble-based systems.

Figure 1

Physical and crystal structures of a typical ZnO NR used in the fluorescence experiment. Approximately 100 individual ZnO NRs were evaulated in the temporal and spatial measurements of biomolecular fluorescence on individual ZnO NRs. (A) Representative BF, DF, and SEM images of a ZnO rod are displayed. The NR in the panels is 1.0 μm and 11.1 μm in diameter and length, respectively. The hexagonal end facets as well as well-defined side facets of a wurtzite ZnO NR are clearly shown in the SEM inset. (B) XRD data of a typical batch of ZnO NRs display predominant 1D growth characteristics along the c-axis of the NRs. The inset clearly displays the low-intensity peaks arising from ZnO NR pyramidal and prismic planes.

Figure 2

Biomolecular fluorescence characteristics observed from the polymeric substrates of PMMA and PS. Black symbols and red lines correspond to the experimental data and curve fit, respectively. (A) The autofluorescence intensities from the polymeric substrates are profiled with respect to the exposure time, t. Both PMMA and PS show non-negligible autofluorescence where the original autofluorescence of PMMA is five times higher than that from PS. (B and C) Normalized, autofluorescence-subtracted fluorescence intensity from 200 μg/ml DTAF-antiIgG on PMMA (B) and on PS (C) is measured at constant illumination and plotted with respect to the exposure time. (D) 500 nm × 500 nm AFM topography panels showing the differences in the adsorbed IgG amount on the PMMA (left) and PS (right) platforms under the same biodeposition condition. Protein adsorption is favored on PS, showing higher amounts of IgG on PS.

Figure 3

Fluorescence intensity profile along the material length and time. (A) The contour maps display variations in the biomolecular fluorescence intensity along the long axis of a 2 μm-long ZnO NR (left) and a 25 μm-long ZnO NR (right). In all cases, fluorescence intensity is enhanced on the basal planes of a ZnO NR. (B) The contour map displays the biomolecular fluorescence intensity over time measured from PS. No significant position-dependent intensity differences are observed. (C) In order to compare the amount of proteins adsorbed on ZnO NR versus PS platforms, UV-vis spectra for the characteristic absorbance peak of an oxidized TMB product at 650 nm are obtained from the two platforms treated identically with 50 μg/ml HRP. The amount of surface-bound HRP is higher on PS than on ZnO NR platforms. (D) Time-lapse images of a typical fluorescence measurement from 200 μg/ml DTAF-antiIgG on a single ZnO NR are displayed.

Figure 4

Biomolecular fluorescence characteristics observed from 200 μg/ml DTAF-antiIgG on a single ZnO NR. Symbols and solid lines in each graph correspond to the experimental data and curve fits, respectively. (A) Differences in the time-dependence of the fluorescence intensity decay under constant irradiation are clearly observed depending on the crystallographic planes within a single ZnO NR. Red and black data show biomolecular fluorescence measured from the basal and prismic planes, respectively. When compared to the signal from the main body of the NR, fluorescence from the NR end is observed not only higher in intensity but also lasts for more prolonged time. (B) The plots show normalized fluorescence intensity profiles for the end (left) and the main body (right) of the ZnO NR, respectively. Each graph is produced by normalizing the two data sets in (A) with regard to the maximum and minimum intensity values of the respective set. (C) Early-time photostability trends of the biomolecular fluorescence measured from the NR basal (red) and the NR prismic (black) planes are displayed in this magnified view of the normalized intensity graphs in (B). Fluorescence signal from the basal plane persists much longer (T_1/2 = 59 sec) than that from the prismic plane (T_1/2 = 40 sec).

Figure 5

FDTD simulations on the fluorescence emission profile from a fluorophore located at 10 nm away from the surface of a ZnO NR. Snap shot images of the three simulation cases for the dipole polarization, (a) parallel to the ZnO NR, (b) in the transverse direction of the NR, and (c) perpendicular to the ZnO NR, are presented. The full movies containing the radiation flow for the three cases are provided in Figure S2†, ESI.