Early diagnosis of oral cancer based on the surface plasmon resonance of gold nanoparticles

Abstract

The high mortality rate in cancer such as oral squamous cell carcinoma is commonly attributed to the difficulties in detecting the disease at an early treatable stage. In this study, we exploited the ability of gold nanoparticles to undergo coupled surface plasmon resonance and set up strong electric fields when closely-spaced to improve the molecular contrast signal in reflectance-based imaging and also to enhance the Raman signal of bioanalytes in cancer. Colloidal gold nanoparticles were synthesized and conjugated to anti-epidermal growth factor receptor (EGFR) for imaging. A self-assembled surface enhanced Raman scattering (SERS)-active gold nanoparticle monolayer film was also developed as a biosensing surface using a simple drop-dry approach. We have shown that gold nanoparticles could elicit an optical contrast to discriminate between cancerous and normal cells and their conjugation with antibodies allowed them to map the expression of relevant biomarkers for molecular imaging under confocal reflectance microscopy. We have also shown that the SERS spectra of saliva from the closely-packed gold nanoparticles films was differentiable between those acquired from normal individuals and oral cancer patients, thus showing promise of a simple SERS-based saliva assay for early diagnosis of oral cancer.

Figure 1

(a) Extinction spectrum of synthesized 15 nm gold nanoparticles with a peak plasmon resonance wavelength of 520 nm and (b) their side-scatter spectrum showing a peak reflectance of the gold nanoparticles at around 600 nm.

Figure 2

Changes in extinction spectrum of gold nanoparticles after conjugation with anti-EGFR to demonstrate the binding of anti-EGFR on gold nanoparticles. Spectrum after conjugation is shown in dashed line.

Figure 3

Confocal reflectance images of CNE2 cells (a) before labeling, (b) after labeling with our control BSA conjugated gold nanoparticles and (c) after labeling with anti-EGFR conjugated gold nanoparticles. Images are cross-sectional slices of cells taken at the mid-focal plane at 20X magnification. False-color reflectance images obtained at excitation 633 nm. Scale bar in all images is 20 μm.

Figure 4

Autofluorescence image of (a) NHLF and (b) CNE2 cells. Their corresponding confocal reflectance images after labeling the (c) NHLF and (d) CNE2 cells with anti-EGFR gold nanoparticles is shown below the autofluorescence image. Images are cross-sectional slices of cells taken at the mid-focal plane at 20X magnification. Falsecolor fluorescence images obtained at excitation 488 nm and reflectance images obtained at excitation 633 nm. Scale bar in all images is 20 μm.

Figure 5

Macroscopic and microscopic images of colloid films by drying 150 μl of 15 nm gold nanoparticles on a SuperFrost^®Plus microscope slide. (a) Macroscopic image of three colloid films. (b) Magnified view showing one fragment of a colloid film.

Figure 6

Normalized UV-Vis spectrum of a gold nanoparticles film obtained by drying a suspension droplet with an initial particle concentration of 1.4 × 10¹² particles per ml (spectrum A) and the 15 nm colloidal gold nanoparticles (spectrum B).

Figure 7

A 1 μm × 1 μm AFM image of the colloid film, showing closely-packed morphology of the gold nanoparticles in the film.

Figure 8

Typical SERS spectra of saliva samples collected from normal healthy individuals (top) and oral-cancer patients (bottom).