Two photon fluorescence sensors based on resonant grating waveguide structures (original) (raw)
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Enhanced two-photon fluorescence excitation by resonant grating waveguide structures
Optics Letters, 2004
Enhanced two-photon f luorescence (TPF) spectroscopy with novel high-finesse resonant polymeric grating waveguide structures (GWSs) is presented. Under resonant conditions the field enhancement at the surface of a GWS can be exploited for TPF spectroscopy without the need for highly focused laser excitation light. We compare the TPF obtained by placing a drop of tetramethylrhodamine (TMR) on top of a GWS with that obtained with TMR on top of a glass substrate. Our procedure and results indicate that the detection of TPF can be improved by a factor of 10 with resonant GWSs.
Applied Physics Letters, 2005
We report a strong enhancement of two-photon fluorescence ͑TPF͒ excitation in the evanescent field of a double grating waveguide structure ͑DGWS͒. For a suitable combination of wavelength, polarization, and angular orientation of the incident laser light DGWSs show resonant behavior resulting in a large field enhancement at the waveguide surface. We demonstrate that at resonance, TPF spectroscopy reveals a 330-fold enhancement of the fluorescence signal of a tetramethylrhodamine thin film prepared from a picomolar aqueous solution. This shows the large potential of DGWSs as TPF-based high-sensitivity sensor platforms for biotechnological and biophysical application.
Giant enhancement of two-photon fluorescence induced by resonant double grating waveguide structures
Applied Physics B, 2004
We report a 350-fold enhancement of ultra-short-pulse-excited twophoton fluorescence (TPF) using a resonant double grating waveguide structure (DGWS). These structures show vanishing transmission and maximum reflection under resonance conditions, i.e. specific wavelength, polarisation and angular orientation of the incident light. This guided mode phenomenon is characterised by a large field enhancement inducing an enormous TPF signal of fluorescent molecules at the waveguide surface, as compared to direct non-resonant excitation. We demonstrate that high spectral acceptance for ultra-short pulses with broad spectral bandwidths can be achieved by a specifically designed DGWS, and that neither beam focussing nor high laser power is necessary for TPF excitation. Due to the high enhancement of more than two orders of magnitude, DGWS can be considered as a powerful platform for TPF applications such as biosensing and microarray technology.
Optics Letters, 2005
We report enhancement of two-photon fluorescence (TPF) excitation in fluorescent dyes and fluorescently labeled biomolecules by exploiting the optical properties of double grating waveguide structures (DGWSs). Picosecond laser pulses generate a large evanescent field based on the guided mode phenomenon in the resonant DGWSs, which induces strong TPF signals from fluorescent dyes at the waveguide surface. By recording enhanced TPF signals of Rhodamine B and Lucifer Yellow under resonance conditions, a detection sensitivity of concentrations of approximately one dye molecule per 0.1 m 2 was achieved. For the first time to our knowledge, enhanced TPF signals of a Lucifer Yellow-labeled biomolecule (human self-peptide) in an aqueous environment are demonstrated. These results strongly encourage the use of DGWSs as enhancement platforms in modern biophysics and biotechnology for investigations of biological membranes and cells.
Selective excitation through tapered silica fibers of fluorescent two-photon polymerized structures
Applied Physics A, 2011
Two-photon polymerization has emerged as a powerful tool to design complex three-dimensional microstructures for applications ranging from biology to nanophotonics. To broaden the application spectrum of such microstructures, different materials have been incorporated to the polymers, aiming at specific applications. In this paper we report the fabrication of microstructures containing rhodamine 610, which display strong fluorescence upon one-and two-photon excitation. The latter increases light-penetration depth and spatial selectivity of luminescence. We also demonstrate that by using silica submicrometric wires we were able to select individual microstructures to be excited, which could be explored for designing microstructure-based optical circuits.
Sensors and Actuators B: Chemical, 2012
Anabolic steroids Methylboldenone Immunosensor Two photon fluorescent spectroscopy (TPF) Double grating waveguide structure (DGWS) Alkyl phosphates and phosphonates SAM Tantalum oxide (Ta2O5) a b s t r a c t An immunosensor approach based on non-linear two-photon fluorescence spectroscopy (TPF) coupled to a resonant double grating waveguide structure (DGWS) has been used for the detection of methylboldenone (MB), an androgenic anabolic steroid used illegally as growth promoter. This synthetic steroid is detected by the biosensor down to 0.1 g l −1 , two orders of magnitude lower than the minimum required performance limit (MRPL) required by the World Anti-Doping Agency (WADA). Nevertheless, we have focused this work on the synthesis and characterization of small fluorescent-labeled conjugates, as well as on the modification of the waveguide surfaces. Detection relies on a direct competitive format, using a boldenone-rhodamine conjugate as fluorescent competitor and a specific anti-MB antibody. The immunoreagents are immobilized onto a resonant Ta 2 O 5 sensing chip after being activated with phosphonohexanoic acid spacers, a novel methodology for covalently surface immobilization of biomolecules, which shows better stability and homogeneity than classical silane chemistries. The developed immunosensor presents great potential as a robust device for its implementation on the detection of small illegal and toxic contaminants.
Two-photon fluorescence excitation of macroscopic areas on planar waveguides
Biosensors and Bioelectronics, 2003
In this paper, we report the first successful demonstration, to our knowledge, of two-photon fluorescence excitation (TPFE) using planar thin-film waveguide structures of macroscopic excitation dimensions (square millimeters to square centimeters in size). The high intensity of excitation light required for TPFE is available not only at a single focus point but along the whole trace of the beam guided in the waveguide structure. Line profiles of the fluorescence excited by TPFE show excellent correlation with the geometry of the launched laser beams. A clear second-order dependence of the fluorescence intensity on the excitation intensity confirms the twophoton character of fluorescence generation. Spectra of the emission generated by one-photon excitation and by two-photon excitation show only minor differences. #
Polymer on Quartz Waveguide Sensing Platform for Enhanced Evanescent Fluorescence Spectroscopy
IEEE Photonics Journal, 2018
This paper presents the study of the performance of a novel SU-8 waveguide on a quartz substrate for evanescent fluorescence spectroscopy. The sensitivity of the sensing platform (SU-8/quartz) was compared to an SU-8 waveguide on silica fabricated with the standard protocol. The physical properties of the SU-8/quartz waveguide resulting from a novel fabrication process allowed for a higher fluorescence coupling and lower optical losses than the SU-8/silica waveguide. The impact of the different indices of refraction of both waveguides on the fluorescence collection efficiency was calculated with three-dimensional finite-difference time-domain simulations by simulating randomly oriented and phased dimensionless current dipoles in the vicinity of their sensing layers. An evanescent fluorescence spectroscopy experiment was performed with different concentrations of Alexa-647 labeled-BSA proteins immobilized on the two polymer waveguides to compare the sensitivity of both sensing platforms. The SU-8/quartz waveguide revealed to have a higher calculated fluorescence collection efficiency and also a greater measured fluorescence light output compared to the SU-8/silica waveguide.
Applied Physics B, 2001
Two-photon excitation (TPE) of fluorescence is a powerful tool for separating a faintly emitted fluorescence signal from background excitation noise. Until now TPE has only been accomplished for very small excitation areas of a few square micrometre dimensions since they are readily available in the focal zone of high-power lasers. In this paper we demonstrate, to our knowledge for the first time, two-photon excited fluorescence with planar thin-film waveguide structures of macroscopic excitation areas of the order of square millimetres to square centimetres.
Journal of Lightwave Technology, 2015
We present a rapid, low-power testbed for the detection and imaging of fluorescent probes utilizing two-photon excitation fluorescence (2PEF). The 2PEF signal from fluorophores commonly used in biological imaging have been enhanced using plasmonic substrates that have been designed to have a high plasmonic resonance over a narrow band that matches the source. The samples were illuminated and imaged using a low-power, in-house multiphoton microscope. A green fluorescent protein (bfloGFPa1), chlorophyll, or rhodamine 6G were deposited onto the plasmonic substrates and their 2PEF signals were measured relative to planar samples. The fluorescence intensities from the green fluorescent protein, chlorophyll, and rhodamine were enhanced by factors of 50, 8, and 4, respectively, with the structured substrates (relative to the planar substrates).