Total Internal Reflection Fluorescence

Axelrod, Daniel; Burghardt, Thomas P.; Thompson, Nancy L.

doi:10.1146/annurev.bb.13.060184.001335

Cited by 646 publications

(346 citation statements)

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“…The dependence of the SF intensity, Isf, on the angular frequency of IR pulses, ωir, may be described by: 31 Isf(ωir) = | χ tot (2) (ωir)| 2 Iir(ωir)Ivis (1) where χ tot (2) is the second-order susceptibility, and Iir and Ivis are the intensities of IR and visible pulses, respectively. The second-order susceptibility can be divided into two components, i.e., the nonlinear susceptibility of gold, χ NR (2) , and the resonant susceptibility, χ R,v (2) , of a vibrational mode, v:…”

Section: Tir-bbsfgmentioning

confidence: 99%

“…For the study of surface phenomena at various interfaces, the total internal reflection (TIR) condition has been used in a number of spectroscopic measurements, 1 such as fluorometry, [2][3][4][5][6][7][8][9] surface enhanced infrared spectroscopy (SEIRA), [10][11][12] second harmonic generation, [13][14][15][16][17][18][19][20][21] and sum frequency generation (SFG). [22][23][24][25] There are two advantages when one uses the TIR condition.…”

Section: Introductionmentioning

confidence: 99%

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Total-internal-reflection Broad-bandwidth Sum Frequency Generation Spectroscopy of Hexadecanethiol Adsorbed on Thin Gold Film Deposited on CaF, 2

et al. 2003

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IntroductionFor the study of surface phenomena at various interfaces, the total internal reflection (TIR) condition has been used in a number of spectroscopic measurements, 1 such as fluorometry, 2-9 surface enhanced infrared spectroscopy (SEIRA), [10][11][12] second harmonic generation, [13][14][15][16][17][18][19][20][21] and sum frequency generation (SFG). [22][23][24][25] There are two advantages when one uses the TIR condition. First, both incident light to the interface and reflected light or generated light from the interface travel in only one of the two phases adjacent to the interface, which circumvents bulk absorption by the other phase. One corollary is the freedom of changing the composition of the adjacent phase that has a lower refractive index. Second, high sensitivity to the interfaces can be achieved owing to the enhancement of the electric field at the interfaces. 1 The TIR arrangement can be employed at metal interfaces if a very thin metal film is used to avoid the absorption of light. It has been successfully employed to detect adsorbed species on metal in the SEIRA, where the thickness of the metal film is about 10 nm on a transparent substrate. [10][11][12] Recently, Williams et al. showed that the TIR arrangement can also be employed to detect SFG signals from alkanethiols adsorbed on a thin gold film. 24 However, detailed analysis of the sum frequency vibrational spectra has not been reported. SFG in the TIR arrangement has some potential as a powerful tool to study molecular orientations at metal/liquid interfaces, such as electrochemical interfaces, because it is free from bulk absorption and has interfacial sensitivity in the order of nanometers. Unlike surface plasmon resonance and capacitance measurements that have been used as sensitive means of detecting adsorption and molecular interaction at the interfaces, TIR-SFG will enable us to obtain molecular information on the solution side of the interfaces. One drawback of the TIR arrangement is the roughness of the metal surfaces. The roughness is prerequisite for high sensitivity and can complicate the interpretation of the experimental data because of the orientational distribution of molecules in comparison with those adsorbed on a single crystal surface. It is therefore of crucial importance to assess the degree of the variation in orientation of molecules.By using broad-bandwidth sum frequency generation (BBSFG) [26][27][28] we recently analyzed the orientation of alkanethiols adsorbed on Au(111). 29It is worthwhile to examine the applicability of BBSFG in the TIR condition at metal interfaces to obtain SFG spectra with higher resolution. In the present paper, we report the results of TIR-BBSFG measurements for hexadecanethiol (HDT) adsorbed on a thin gold film on CaF2. We will show that BBSFG can be successfully employed in the orientation analysis of the HDT molecules and that the methyl groups of HDT are much more randomly orientated than those on Au(111) on mica. Experimental SubstrateA hemicylindrical CaF2 prism (Pier Optic...

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Section: Tir-bbsfgmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Total-internal-reflection Broad-bandwidth Sum Frequency Generation Spectroscopy of Hexadecanethiol Adsorbed on Thin Gold Film Deposited on CaF, 2

et al. 2003

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“…1 shows the coordinate system in which the excitation beam is reflected at the interface between the silica slide with a higher optical density, n 1 , and the liquid, n 2 . The electric field amplitude of the evanescent wave, E, decreases exponentially in solution (z-direction) (27):…”

Section: Calibration Of Tirfmentioning

confidence: 99%

Adsorption Kinetics, Conformation, and Mobility of the Growth Hormone and Lysozyme on Solid Surfaces, Studied with TIRF

Buijs

Hlady

1997

Journal of Colloid and Interface Science

158

122

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Interactions of recombinant human growth hormone and lysozyme with solid surfaces are studied using total internal reflection fluorescence (TIRF) and monitoring the protein's intrinsic tryptophan fluorescence. The intensity, spectra, quenching, and polarization of the fluorescence emitted by the adsorbed proteins are monitored and related to adsorption kinetics, protein conformation, and fluorophore rotational mobility. To study the influence of electrostatic and hydrophobic interactions on the adsorption process, three sorbent surfaces are used which differ in charge and hydrophobicity. The chemical surface groups are silanol, methyl, and quaternary amine. Results indicate that adsorption of hGH is dominated by hydrophobic interactions. Lysozyme adsoption is strongly affected by the ionic strength. This effect is probably caused by an ionic strength dependent conformational state in solution which, in turn, influences the affinity for adsorption. Both proteins are more strongly bound to hydrophobic surfaces and this strong interaction is accompanied by a less compact conformation. Furthermore, it was seen that regardless of the characteristics of the sorbent surface, the rotational mobility of both proteins' tryptophans is largely reduced upon adsorption.

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“…While analytical methods such as fluorescence correlation spectroscopy (FCS) 1 and fluorescence recovery after photobleaching 2 can be used with diffraction-limited optical systems, there exist several approaches for beating the diffraction limit. Total internal reflection fluorescent microscopy (TIRF) 3 and near field scanning optical microscopy 4,5 rely on decaying evanescent waves to create a subdiffraction limited focal volume. Twophoton excited fluorescence is also available as a method for shrinking illumination volume and increasing signal-tonoise, 6,7 and can be used noninvasively on living organisms.…”

Section: Introductionmentioning

confidence: 99%

Nanofluidic structures for single biomolecule fluorescent detection

Mannion

Craighead

2006

Biopolymers

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Fluid‐filled nanofabricated cavities can be used to increase the spatial resolution of single molecule confocal microscopy based techniques by creating smaller and more uniformly illuminated probe volumes. Such structures may also be used to temporarily stretch single macromolecules, permitting the resolution of molecular details that would otherwise be beyond the capabilities of a diffraction limited system. © 2006 Wiley Periodicals, Inc. Biopolymers 85:131–143, 2007.This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com

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Total Internal Reflection Fluorescence

Cited by 646 publications

References 68 publications

Total-internal-reflection Broad-bandwidth Sum Frequency Generation Spectroscopy of Hexadecanethiol Adsorbed on Thin Gold Film Deposited on CaF, 2

Total-internal-reflection Broad-bandwidth Sum Frequency Generation Spectroscopy of Hexadecanethiol Adsorbed on Thin Gold Film Deposited on CaF, 2

Adsorption Kinetics, Conformation, and Mobility of the Growth Hormone and Lysozyme on Solid Surfaces, Studied with TIRF

Nanofluidic structures for single biomolecule fluorescent detection

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