Photonic crystal enhanced fluorescence using a quartz substrate to reduce limits of detection

Abstract

A Photonic Crystal (PC) surface fabricated upon a quartz substrate using nanoimprint lithography has been demonstrated to enhance light emission from fluorescent molecules in close proximity to the PC surface. Quartz was selected for its low autofluorescence characteristics compared to polymer-based PCs, improving the detection sensitivity and signal-to-noise ratio (SNR) of PC Enhanced Fluorescence (PCEF). Nanoimprint lithography enables economical fabrication of the subwavelength PCEF surface structure over entire 1x3 in2 quartz slides. The demonstrated PCEF surface supports a transverse magnetic (TM) resonant mode at a wavelength of λ = 632.8 nm and an incident angle of θ = 11°, which amplifies the electric field magnitude experienced by surface-bound fluorophores. Meanwhile, another TM mode at a wavelength of λ = 690 nm and incident angle of θ = 0° efficiently directs the fluorescent emission toward the detection optics. An enhancement factor as high as 7500 × was achieved for the detection of LD-700 dye spin-coated upon the PC, compared to detecting the same material on an unpatterned glass surface. The detection of spotted Alexa-647 labeled polypeptide on the PC exhibits a 330 × SNR improvement. Using dose-response characterization of deposited fluorophore-tagged protein spots, the PCEF surface demonstrated a 140 × lower limit of detection compared to a conventional glass substrate.

Fig. 1

Schematic diagram of PCEF surface on a quartz substrate. The subwavelength grating structure is etched into the quartz substrate and a high refractive index dielectric (TiO₂) film is coated on top of the grating as a light confinement layer.

Fig. 2

(a) RCWA simulated dispersion diagram for the PC used in this study. Resonance for the enhanced excitation for the TM mode is at ~10.8°(b) Simulated near field distribution at λ = 632.8 nm (normalized to the intensity of the incident field).

Fig. 3

Schematic diagram of the fabrication procedure: (a) The process begins with a dispense pattern of MonoMat on a planarized quartz wafer; (b) The template is pressed against the dispense pattern and then UV cured; (c) The template is pulled away from the solidified grating pattern; (d) A layer of Silspin is spin coated onto the patterned surface; (e) RIE of SilSpin to expose imprint resist; (f) RIE of the imprint resist to expose the quartz surface; (g) RIE of quartz to transfer the pattern onto the wafer; (h) Piranha cleaning of the wafer to remove the imprint resist residues; (i) TiO₂ deposition onto the grating.

Fig. 4

(a) SEM image of the top view of the TiO₂ coated grating structure on quartz substrate; (b) AFM image of the PCEF surface showing the grating depth of 40 nm; (c) Photograph of the PCEF surface on 1 × 3 in² substrate.

Fig. 5

(a) Wavelength transmission spectrum; (b) Angle transmission spectrum at the excitation wavelength λ = 632.8nm.

Fig. 6

Autofluorescne intensity from a normal glass slide, a plastic-based PCEF surface, and a quartz-based PCEF surface measured using the PC fluorescent microscope under identical measurement settings.

Fig. 7

Fluorescence output as a function of angle of incidence for a ~50 nm film of dye-doped polymer applied directly to the PCEF surface.

Fig. 8

Angle-resolved fluorescence measurement on the quartz PCEF surface.

Fig. 9

(a) Gain and exposure-optimized images of PPL-Alexa 647 fluorescence on glass compared the PCEF surface; (b) Intensity profile as a function of distance for line of fluorescent image pixels profiling spots of concentration 9.9 µg/ml on both glass and the PCEF surface.

Fig. 10

Signal-to-noise vs PPL-Alexa 647 concentration showing an improvement in limit of detection (LOD) on a PCEF surface by a factor of 140.