Detection and Quantification of DNA by Fluorophore-Induced Plasmonic Current: A Novel Sensing Approach

Abstract

We report on the detection and quantification of aqueous DNA by a fluorophore-induced plasmonic current (FIPC) sensing method. FIPC is a mechanism described by our group in the literature where a fluorophore in close proximity to a plasmonically active metal nanoparticle film (MNF) is able to couple with it, when in an excited state. This coupling produces enhanced fluorescent intensity from the fluorophore-MNF complex, and if conditions are met, a current is generated in the film that is intrinsically linked to the properties of the fluorophore in the complex. The magnitude of this induced current is related to the spectral properties of the film, the overlap between these film properties and those of the fluorophore, the spacing between the nanoparticles in the film, the excitation wavelength, and the polarization of the excitation source. Recent literature has shown that the FIPC system is ideal for aqueous ion sensing using turn-on fluorescent probes, and in this paper, we subsequently examine if it is possible to detect aqueous DNA also via a turn-on fluorescent probe, as well as other commercially available DNA detection strategies. We report the effects of DNA concentration, probe concentration, and probe characteristics on the development of an FIPC assay for the detection of non-specific DNA in aqueous solutions.

Keywords: DNA sensing; fluorophore induced plasmonic current; metal-enhanced fluorescence; plasmonic current; plasmonic electricity.

Figure 1

Absorption spectra of various solutions of ethidium bromide, ranging from 20 µM to 100 µM, both with and without the addition of 1 mg/mL DNA from a salmon sperm DNA stock solution. The solutions were allowed to mix for 30 min prior to analysis.

Figure 2

Fluorescence emission spectra of various solutions of ethidium bromide, ranging from 20 µM to 100 µM, both with and without the addition of 1 mg/mL DNA from a salmon sperm DNA stock solution, excited at 365 nm. The solutions were allowed to mix for 30 min prior to analysis.

Figure 3

Peak fluorescence emission spectra values for various solutions of ethidium bromide, ranging from 20 µM to 100 µM with the addition of 1 mg/mL DNA from a salmon sperm DNA stock solution, excited at 365 nm. The solutions were allowed to mix for 30 min prior to analysis. Data organized as the peak fluorescence intensity at 600 nm vs. concentration.

Figure 4

Plasmonic current response of various solutions of ethidium bromide, ranging from 20 µM to 100 µM, both with and without the addition of 1 mg/mL DNA from a salmon sperm DNA stock solution, excited at 266 nm at 100 µW excitation power. The solutions were allowed to mix for 30 min prior to analysis.

Figure 5

Plasmonic current response of various solutions of ethidium bromide, ranging from 20 µM to 100 µM with the addition of 1 mg/mL DNA from a salmon sperm DNA stock solution, excited at 266 nm at 100 µW excitation power. The solutions were allowed to mix for 30 min prior to analysis.

Figure 6

Absorbance spectra of a 50µM solution of ethidium bromide mixed with varying concentrations of DNA from a salmon sperm DNA stock solution, ranging from 1 µg/mL to 10,000 µg/mL. The solutions were allowed to mix for 30 min prior to analysis.

Figure 7

Fluorescence emission spectra of a 50 µM of ethidium bromide mixed with varying concentrations of DNA from a salmon sperm DNA stock solution, ranging from 1 µg/mL to 10,000 µg/mL, excited at 266 nm. The solutions were allowed to mix for 30 min prior to analysis.

Figure 8

Peak fluorescence emission spectra values of a 50 µM of ethidium bromide solution mixed with varying concentrations of DNA from a salmon sperm DNA stock solution, ranging from 1 µg/mL to 10,000 µg/mL, excited at 266 nm. The solutions were allowed to mix for 30 min prior to analysis. Data shown as peak fluorescence intensity at 600 nm vs. DNA concentration.

Figure 9

Plasmonic current response of a 50 µM of ethidium bromide solution mixed with varying concentrations of DNA from a salmon sperm DNA stock solution, ranging from 1 µg/mL to 10,000 µg/mL, excited at 266 nm at 100 µW power. The solutions were allowed to mix for 30 min prior to analysis.

Figure 10

Absorbance spectra of various solutions of SYBR Green with varying concentrations of added DNA. Concentrations in μg/mL and mg/mL are related to DNA concentration added; 100×, 10× and 1× are related to the concentrations of SYBR Green.

Figure 11

Fluorescence spectra of various solutions of SYBR Green with different concentrations of DNA added, excited at 473 nm. (A) 1× Concentration, (B) 10× Concentration and, (C) 100× Concentration of SYBR Green.

Figure 12

FIPC responses for various solutions of SYBR Green with various amounts of DNA added, excited via a 473 nm (CW) laser, 10 mW power.

Figure 13

FIPC response for 1× SYBR Green with added DNA, excited via 473 nm laser. Comparison between P and S polarized light for excitation.

Figure 14

Standard curve obtained using 1× SYBR Green and various DNA concentrations (Left). In a blinded study, the sample values were found to be within 15% of their true value. Table detailing the raw responses and their calculated values (on the right). Each current response shown is the average of 10 values, n = 10.