Single Molecule Detection by Laser Two-Photon Excited Fluorescence in a Capillary Flowing Cell

Song, Joon Myong; Inoue, Takanori; Kawazumi, Hirofumi; Ogawa, Teiichiro

doi:10.2116/analsci.14.913

Cited by 10 publications

(8 citation statements)

References 18 publications

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“…They observed photon bursts from single molecules diffusing through the two-photon volume, with an average SBR of ∼10. A number of other reports have since demonstrated efficient fluorescence burst detection of single molecules by TPE in free solution (72,73), in flow cells (74)(75)(76), and in low-temperature solids (77,78). As anticipated, a common finding among most of these reports is the high SBR, largely caused by the very low background levels associated with TPE.…”

Section: Single-molecule Detection In Solutionmentioning

confidence: 85%

Two-Photon Excitation Fluorescence Microscopy

Chen

Masters

et al. 2000

Annu. Rev. Biomed. Eng.

939

507

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Two-photon fluorescence microscopy is one of the most important recent inventions in biological imaging. This technology enables noninvasive study of biological specimens in three dimensions with submicrometer resolution. Two-photon excitation of fluorophores results from the simultaneous absorption of two photons. This excitation process has a number of unique advantages, such as reduced specimen photodamage and enhanced penetration depth. It also produces higher-contrast images and is a novel method to trigger localized photochemical reactions. Two-photon microscopy continues to find an increasing number of applications in biology and medicine.

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Section: Single-molecule Detection In Solutionmentioning

confidence: 85%

Two-Photon Excitation Fluorescence Microscopy

Chen

Masters

et al. 2000

Annu. Rev. Biomed. Eng.

939

507

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“…to 1PA-PL 1,4,15 and has emerged as an important nonlinear optical detection technique, finding applications such as single molecule/nanoparticle detection 16 , fluorescence correlation spectroscopy 17,18 and multi-photon high contrast bio imaging [19][20][21] . Such 2PA-PL probed by high intensity ultrashort pulses reveals information from much deeper depths of the sample and several underlying effects [20][21][22] (such as lattice defects and layered misalignments) and new excited state relaxations.…”

mentioning

confidence: 99%

Linear and nonlinear optical probing of various excitons in 2D inorganic-organic hybrid structures

Adnan

Baumberg

Prakash

2020

Sci Rep

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Nonlinear optical properties, such as two-(or multi-) photon absorption (2PA), are of special interest for technologically important applications in fast optical switching, in vivo imaging and so on. Highly intense infrared ultrashort pulses probe deep into samples and reveal several underlying structural perturbations (inter-layer distortions, intra-layer crumpling) and also provide information about new excited states and their relaxation. naturally self-assembled inorganic-organic multiple quantum wells (io-MQWs) show utility from room-temperature exciton emission features (binding energies ~200-250 meV). These Mott type excitons are highly sensitive to the self-assembly process, inorganic network distortions, thickness and inter-layer distortions of these soft two-dimensional (2D) and weak van der Waal layered hybrids. We demonstrate strong room-temperature nonlinear excitation intensity dependent two-photon absorption induced exciton photoluminescence (2PA-PL) from these IO-MQWs, excited by infrared femtosecond laser pulses. Strongly confined excitons show distinctly different one-and two-photon excited photoluminescence energies: from free-excitons (2.41 eV) coupled to the perfectly aligned MQWs and from energy down-shifted excitons (2.33 eV) that originate from the locally crumpled layered architecture. High intensity femtosecond induced pL from one-photon absorption (1PA-PL) suggests saturation of absorption and exciton-exciton annihilation, with typical reduction in PL radiative relaxation times from 270 ps to 190 ps upon increasing excitation intensities. From a wide range of IR excitation tuning, the origin of 2PA-PL excitation is suggested to arise from exciton dark states which extend below the bandgap. Observed two-photon absorption coefficients (β ~75 cm/ GW) and two-photon excitation cross-sections (η 2 σ 2 ~ 110GM), further support the evidence for 2PA excitation origin. Both 1PA-and 2PA-PL spatial mappings over large areas of single crystal platelets demonstrate the coexistence of both free and deep-level crumpled excitons with some traces of defectinduced trap state emission. We conclude that the two-photon absorption induced pL is highly sensitive to the self-assembly process of few to many mono layers, the crystal packing and deep level defects. This study paves a way to tailor the nonlinear properties of many 2D material classes. Our results thus open new avenues for exploring fundamental phenomena and novel optoelectronic applications using layered inorganic-organic and other metal organic frameworks. High intensity ultrashort laser pulses interact with nanomaterials to produce fascinating parametric and nonparametric nonlinear optical phenomena such as two-photon (or multi-photon) absorption 1 , harmonic generation 2 , coherent anti-Stokes Raman scattering (CARS) 3 , excited state absorption and ultrafast charge carrier dynamics 4,5. Studies of two-(or multi-) photon absorption in layered materials (such as graphene oxide, MoS 2 type transition-metal dichalcogenides) 6-11 have given new in...

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“…3 In brief, a self modelocked Ti-sappihire laser (Coherent Mira 900; pulsed width 180 fs, repetition rate 76 MHz) was tuned to 800 nm and focused on a BBO crystal for second harmonic generation (400 nm). The average power of the fundamental beam and the second harmonic beam was measured as 0.27 W and 0.10 W, respectively.…”

Section: Methodsmentioning

confidence: 99%

“…1 Highly sensitive determinations based on two-photon excited fluorescence have been carried out using a high-intensity laser. 2,3 Characteristics of two-photon absorption should be clarified for better understanding of its mechanism and for developing wider analytical applications. There are several techniques to determine a two-photon absorption cross section.…”

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confidence: 99%

Determination of Two Photon Absorption Cross Section of Fluorescein Using a Mode Locked Titanium Sapphire Laser

et al. 1999

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Laser two-photon excited fluorescence has its own advantages in selection rule and backgrounds over onephoton excited (conventional) fluorescence.1 Highly sensitive determinations based on two-photon excited fluorescence have been carried out using a high-intensity laser.2,3 Characteristics of two-photon absorption should be clarified for better understanding of its mechanism and for developing wider analytical applications. There are several techniques to determine a two-photon absorption cross section. [4][5][6] However, its direct measurement is still difficult because of its small value. 7Due to its low efficiency a sample in a high concentration had to be prepared for a determination of the twophoton absorption cross section, and the obtained value may be susceptible to self absorption. 6 In this report, the two-photon absorption cross section of fluorescein at 800 nm has been determined by measuring the intensity ratio of one-photon excited fluorescence to two-photon excited fluorescence using a mode locked Ti-sapphire laser. Because of the use of the second harmonic of this laser as a light source of one-photon excitation, the present method is theoretically clearer and experimentally simpler than the other methods so far used. This report shows that the high peak power of the Ti:sapphire laser is more suitable for a determination of two-photon absorption cross section than a CW laser. ExperimentalThe apparatus is essentially identical with that described in the previous paper. 3 In brief, a self modelocked Ti-sappihire laser (Coherent Mira 900; pulsed width 180 fs, repetition rate 76 MHz) was tuned to 800 nm and focused on a BBO crystal for second harmonic generation (400 nm). The average power of the fundamental beam and the second harmonic beam was measured as 0.27 W and 0.10 W, respectively. Both of the beams were focused on a capillary cell (J&W Scientific, 100 µm i.d.) using a 10-X microscope objective. The polyimide coating of the capillary was removed at the irradiation position of the laser beam. The fluorescence was collected using another 10-X microscope objective which was set perpendicularly to the incident laser beam so that one-photon and twophoton excited fluorescence could be compared under identical experimental conditions. The fluorescence was transmitted through two glass filters (Andover Y-44: short cut-off below 410 nm; Andover 040FG11: band path at 320 -680 nm and transmission at 800 nm is below 10 -5 ) and was measured with a photomultiplier (Hamamatsu R5600U, quantum efficiency at 800 nm: <10 -5 ). The photocurrent was amplified and counted by a timer and counter (EG&G Ortec 871: 100 MHz).The stock solution of fluorescein (1×10 -4 M) was prepared in 20 mM phosphate buffer at pH 11. The sample solution with 1×10 -6 M was prepared by a successive dilution of the stock solution and injected into the capillary cell by vacuum flushing. Results and Discussion Formulation and fluorescence intensityThe two-photon excited process is less efficient and has not been used much for highly sens...

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Single Molecule Detection by Laser Two-Photon Excited Fluorescence in a Capillary Flowing Cell

Cited by 10 publications

References 18 publications

Two-Photon Excitation Fluorescence Microscopy

Two-Photon Excitation Fluorescence Microscopy

Linear and nonlinear optical probing of various excitons in 2D inorganic-organic hybrid structures

Determination of Two Photon Absorption Cross Section of Fluorescein Using a Mode Locked Titanium Sapphire Laser

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