Two-photon Excited Fluorescence Emission Research Articles

Molecular architecture effects in two-photon absorption: from octupolar molecules to polymers and hybrid polymer nanoparticles based on 1,3,5-triazine.

The two-photon absorption properties of a set of linear copolymers based on the regular alternation of a 2,4,6-tris(thiophen-2-yl)-1,3,5-triazine electron-accepting unit with different electron-donating groups attached to two of the thiophen ends were investigated. Comparison of these data with those of the analogous octupolar monomers and hyperbranched polymers allows us to understand the role of the triazine-thiophen core and its molecular architecture in the nonlinear optical properties of these polymeric materials. It is concluded that the arrangement of the push-pull unit into a unidimensional array, as it is the case of the linear copolymer, favours the two-photon absorption cross-section. Hybrid nanoparticles dispersed in water were prepared from selected polymers with two-photon excited fluorescence emission comparable with those of the best performing quantum dots.

In vivo monitoring of protein-bound and free NADH during ischemia by nonlinear spectral imaging microscopy

Nonlinear spectral imaging microscopy (NSIM) allows simultaneous morphological and spectroscopic investigation of intercellular events within living animals. In this study we used NSIM for in vivo time-lapse in-depth spectral imaging and monitoring of protein-bound and free reduced nicotinamide adenine dinucleotide (NADH) in mouse keratinocytes following total acute ischemia for 3.3 h at ~3 min time intervals. The high spectral resolution of NSIM images allows discrimination between the two-photon excited fluorescence emission of protein-bound and free NAD(P)H by applying linear spectral unmixing to the spectral image data. Results reveal the difference in the dynamic response between protein-bound and free NAD(P)H to ischemia-induced hypoxia/anoxia. Our results demonstrate the capability of nonlinear spectral imaging microscopy in unraveling dynamic cellular metabolic events within living animals for long periods of time.

A cationic triarylborane as water-tolerant fluorescent chemosensor for fluoride anions

A new cationic triarylborane has been synthesized by methylation of a dimethylamino electron donor group in a neutral donor-π-acceptor type system, which also includes a dimesitylboryl group as electron acceptor. Its single- and two-photon excited fluorescence emission properties have been examined, and it has been found to be a highly fluorescent chemosensor for fluoride anions in THF solutions. This cation exhibits a higher sensitivity for fluoride anions compared to its neutral precursor, which is attributed to the increased Lewis acidity of the boron center, as well as its enhanced Coulombic attraction for anions. It has also been demonstrated to be water-tolerant.

Raman-fluorescence hybrid microspectroscopy of cell nuclei

Raman-fluorescence hybrid confocal microspectroscopy of cell nuclei in different stages of nuclear development will be presented. Two-photon excited fluorescence microscopy enables fast detection of the morphology of the nucleus after staining with nucleotide sensitive dyes. Areas of interest are selected from the fluorescence image and subsequently analyzed with chemically selective Raman microspectroscopy. A single excitation source at 647.1 nm from a krypton-ion laser avoids spectral overlap between the Raman spectrum on the Stokes side and the two-photon excited fluorescence emission at the anti-Stokes side. Nucleotides are stained with DAPI or Hoechst 33342. The two-photon response of Hoechst 33342 is higher than that of DAPI. Raman microspectroscopy supports the finding that areas in live cell nuclei with low staining of Hoechst 33342 contain relatively high levels of RNA.

Influence of Polymer Structures on Optical Power Limiting Performance of Single-Walled Carbon Nanotubes

Purified single-walled carbon nanotubes are dispersed in two different types of polymer matrices. In these stable dispersed systems, the interaction between the polymer matrix and the single-walled carbon nanotubes, optical power limiting effect, and two-photon absorption are investigated. Transmission electron micrographs show the rigid polymer backbone interacts with the single-walled carbon nanotubes and results in the formation of nanoribbons and nanobundles in the dispersions. Optical measurements show that the nanoribbons and nanobundles significantly improve the optical power limiting performance through increased nonlinear optical absorption. Detailed comparison of optical power limiting, two-photon absorption, and two-photon excited fluorescence emission is presented. Our experimental results show that the different polymer backbones have a considerable effect on the nonlinear optical properties of carbon nanotubes.

Side illumination fluorescence emission characteristics from a dye doped polymer optical fiber under two-photon excitation

Two-photon excited (TPE) side illumination fluorescence studies in a Rh6G-RhB dye mixture doped polymer optical fiber (POF) and the effect of energy transfer on the attenuation coefficient is reported. The dye doped POF is pumped sideways using 800 nm, 70 fs laser pulses from a Ti:sapphire laser, and the TPE fluorescence emission is collected from the end of the fiber for different propagation distances. The fluorescence intensity of RhB doped POF is enhanced in the presence of Rh6G as a result of energy transfer from Rh6G to RhB. Because of the reabsorption and reemission process in dye molecules, an effective energy transfer is observed from the shorter wavelength part of the fluorescence spectrum to the longer wavelength part as the propagation distance is increased in dye doped POF. An energy transfer coefficient is found to be higher at shorter propagation distances compared to longer distances. A TPE fluorescence signal is used to characterize the optical attenuation coefficient in dye doped POF. The attenuation coefficient decreases at longer propagation distances due to the reabsorption and reemission process taking place within the dye doped fiber as the propagation distance is increased.

Strong two-photon absorption and two-photon excited fluorescence emission of heterofluorene derivatives

Abstract Two new chromophores: 2,8-bis[2-(4′-ethoxylphenyl)-5-(4′-styryl)-1,3,4-oxadiazole]dibenzothiophene (abbreviated as SO-G1) and 2,8-bis[(2-(4′-ethoxylphenyl)-5-(4′-styryl)-1,3,4-oxadiazole- N -ethylcarbazole (abbreviated as NO-G1) that exhibit strong two-photon absorption and enhanced two-photon excited fluorescence were synthesized based on “push–pull–core–pull–push” elongated molecules built from a rigid dibenzothiophene or carbazole. Both chromophores in different solvent are highly transparent across almost the whole visible spectrum region and possess highest fluorescence quantum yield of 0.89 for NO-G1 and 0.46 for SO-G1. Pumped by a visible wavelength (700–800 nm) of Ti: sapphire femto-second laser pulses, strong two-photon excited (TPE) fluorescence located at 440–510 nm with the TPE fluorescence emission cross-section ( δ TPF ) being 21–8.2 times as much as that of fluorescein reference and strong two-photon absorption (TPA) cross-section ( δ TPA ) of 454–377 GM have been observed.

Highly sensitive detection using laser two-photon excited fluorescence in capillary electrophoresis

The laser two-photon excited fluorescence (LTPF) has been applied to a capillary electrophoresis (CE) system using a self mode-locked titanium-sapphire laser. The analytes were coumarine 440 (7-amino-4-methyl coumarine) and DCCS (7-diethyl aminocoumarine-3-carboxylic acid succinimidyl ester). The two-photon excited fluorescence emission spectrum of DCCS was identical to the one-photon excited fluorescence emission spectrum. The LTPF quantum efficiency increased through the addition of methanol to the buffer solution, and, as a result, the detection limit decreased. The detection limit of coumarine 440 was 8.0 amol, this value was as low as that obtained by one-photon excited fluorescence.