SEARCH · PubMed Health

Results for “Microscopy, Fluorescence, Multiphoton”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 397 records · Page 22Linked to original sources

PubMed2004

Enhanced two-photon fluorescence excitation by resonant grating waveguide structures.

Enhanced two-photon fluorescence (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.

Equipment Design↗

PubMed2004

Second-harmonic microscopy of unstained living cardiac myocytes: measurements of sarcomere length with 20-nm accuracy.

We extend second-harmonic generation (SHG) microscopy to the measurement of sarcomere length in unstained living cardiac myocytes with 20-nm accuracy. We quantify individual sarcomere shortening in the presence of saxitoxin and find that it is in agreement with mechanical measurements of atrial tissue contracture. This functional application of SHG microscopy is generally applicable to quantify the physiological effects of drugs on contractile tissue. Our data also suggest that packed myosin heads in sarcomere thick filaments are responsible for the large second-harmonic endogenous signal in muscle tissue.

Animals↗

PubMed2004

Miniaturized two-photon microscope based on a flexible coherent fiber bundle and a gradient-index lens objective.

We present a miniature, flexible two-photon microscope consisting of a fused coherent optical fiber bundle with 30,000 cores and a gradient-index lens objective. The laser focus of a standard two-photon laser-scanning microscope was scanned over the entrance surface of the fiber bundle, resulting in sequential coupling into individual cores. Fluorescent light was detected through the fiber bundle. Micrometer-sized fluorescent beads and pollen grains were readily resolved. In addition, fluorescently labeled blood vessels were imaged through the fiber bundle in rat brain in vivo.

Animals↗

PubMed2005

Fourier transform measurement of two-photon excitation spectra: applications to microscopy and optimal control.

We report a novel Fourier transform method for measuring two-photon excitation spectra. We demonstrate this method using simple dye molecules and discuss its applications in two-photon fluorescence microscopy and optimal control. This method facilitates an intuitive interpretation of recent control experiments in terms of tuning the nonlinear spectrum of the exciting laser source.

Equipment Design↗

PubMed2005

Direct four-photon excitation of amplified spontaneous emission in a nonlinear organic chromophore.

We report four-photon pumped amplified spontaneous emission in an organic chromophore. The phenomenon is observed when intense infrared laser radiation illuminates a nonlinear chromophore solution. As a result of this illumination, a strong and highly directional radiation in the visible wavelength range is generated in both the forward and the backward directions, with an angular divergence similar to that of the pump beam.

Fluorescent Dyes↗

PubMed2005

Two-photon absorption and self-phase modulation measurements with shaped femtosecond laser pulses.

We show that phase-sensitive detection of spectral hole refilling can yield information about self-phase modulation and two-photon absorption coefficients. We expect that, when applied to tissue microscopy, this technique will allow the study of endogenous molecular markers beneath the surface, even if those markers are nonfluorescent.

Algorithms↗

PubMed2005

Picosecond-pulse-induced two-photon fluorescence enhancement in biological material by application of grating waveguide structures.

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 microm2 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.

Equipment Design↗

PubMed2005

Four-color fluorescence correlation spectroscopy realized in a grating-based detection platform.

We have developed a filterless multicolor detection unit for fluorescence correlation spectroscopy (FCS). This grating-based setup is continuously tunable for multicolor separation and is thus a powerful alternative to the classical cascade of dichroic mirrors and filters. Our tailored platform allows for accommodation of up to 15 detection channels covering the entire visible spectral range. As a proof of principle, we successfully demonstrate simultaneous FCS of four distinct fluorescent quantum dot species being mixed in aqueous solution. Grating-based detection allows for spectral high-resolution FCS in a stable and compact setup and is a feasible tool for quantitative investigation of complexbiomolecular dynamics on a single molecule level.

Equipment Design↗

PubMed2005

In vivo brain imaging using a portable 3.9 gram two-photon fluorescence microendoscope.

We introduce a compact two-photon fluorescence microendoscope based on a compound gradient refractive index endoscope probe, a DC micromotor for remote adjustment of the image plane, and a flexible photonic bandgap fiber for near distortion-free delivery of ultrashort excitation pulses. The imaging head has a mass of only 3.9 g and provides micrometer-scale resolution. We used portable two-photon microendoscopy to visualize hippocampal blood vessels in the brains of live mice.

Animals↗

PubMed2005

Three-photon induced photobleaching in a three-dimensional memory material.

The photobleaching properties of a 3D memory material based on pyrylium salts are studied. It is shown that under ultrashort pulsed irradiation photobleaching occurs through two diverse mechanisms. One is major and causes fast bleaching, whereas the other is minor and causes slow bleaching. Furthermore, it is proved that both mechanisms are three- or more-photon processes. This fact gives the capability of significantly increasing data storage density.

Computers, Molecular↗

PubMed2006

Infrared two-photon-excited visible lasing from a DNA-surfactant-chromophore complex.

Infrared two-photon-pumped and cavity-enhanced frequency upconversion lasing has been achieved in a novel DNA-surfactant-chromophore complex (DSCC) gel system, which is a new step toward producing a biological laser. Once the focused intensity of the 150 fs and approximately 775 nm pump laser beam is higher than a certain threshold level, highly directional stimulated emission at approximately 582 nm wavelength can be observed from a 1 cm long DSCC complex gel cell. With cavity feedback provided by the two optical windows, the pump threshold can be further reduced, the highly directional output lasing can be greatly enhanced, and the output spectral linewidth can be reduced to less than 1/5 of the spontaneous fluorescence spectral bandwidth.

DNA↗

PubMed2006

Biomolecular imaging based on far-red fluorescent protein with a high two-photon excitation action cross section.

Received October 14, 2005; revised January 7, 2006; accepted January 9, 2006; posted January 12, 2006 (Doc. ID 65391) The two-photon excitation action cross section of Hc-Red fluorescent proteins (Hc-RFPs) is measured and found to be of the same order as that of enhanced green fluorescent proteins. With a 618 nm emission wavelength in the far-red region and with an excitation wavelength around 1200 nm, Hc-RPF-based two-photon fluorescence microscopy (2PFM) can offer deep penetration capability inside live samples and is ideal for in vivo gene expression study and biomolecular imaging in live objects. In vivo 2PFM of the developing heart deep inside a transgenic zebrafish embryo tagged by Hc-RFP is also successfully demonstrated.

Animals↗

PubMed2006

Fiber-optic scanning two-photon fluorescence endoscope.

We report on the development of a miniature, flexible, fiber-optic scanning endoscope for two-photon fluorescence imaging. The endoscope uses a tubular piezoelectric actuator for achieving two-dimensional beam scanning and a double-clad fiber for delivery of the excitation light and collection of two-photon fluorescence. Real-time imaging of fluorescent beads and cancer cells has been performed.

Endoscopes↗

PubMed2006

Prism-based multicolor fluorescence correlation spectrometer.

We report the design and application of a prism-based detection system for fluorescence (cross) correlation spectroscopy. The system utilizes a single laser wavelength for the simultaneous excitation of several dyes of different emission spectra. Fluorescence light is spectrally separated with a prismatic setup, and wavelengths are selected by scanning a fiber-coupled avalanche photodiode across the image spots. Multicolor autocorrelations are demonstrated with standard and tandem dyes, and fluorescence cross-correlation measurements of biotinylated nanocontainers and streptavidin are presented. This spectrometer offers high optical stability and no focal volume mismatch for the multicolor detection of molecular dynamics and interactions, with single-molecule sensitivity.

Equipment Design↗

PubMed2006

Fast-scanning two-photon fluorescence imaging based on a microelectromechanical systems two- dimensional scanning mirror.

Towards overcoming the size limitations of conventional two-photon fluorescence microscopy, we introduce two-photon imaging based on microelectromechanical systems (MEMS) scanners. Single crystalline silicon scanning mirrors that are 0.75 mm x 0.75 mm in size and driven in two dimensions by microfabricated vertical comb electrostatic actuators can provide optical deflection angles through a range of approximately16 degrees . Using such scanners we demonstrated two-photon microscopy and microendoscopy with fast-axis acquisition rates up to 3.52 kHz.

Computer Systems↗

PubMed2006

Multidimensional single-molecule imaging in live cells using total-internal-reflection fluorescence microscopy.

We have developed a wide-field total-internal-reflection fluorescence microscope capable of imaging single molecules in live cells, resolved in both wavelength and polarization. We show fluorescence resonance energy transfer between single pairs of fluorescent molecules bound to signaling receptors in the plasma membrane of live cells and demonstrate the importance of polarization discrimination in addition to wavelength separation.

Animals↗

PubMed2006

Dual-pump coherent anti-Stokes-Raman scattering microscopy.

We introduce dual-pump coherent anti-Stokes-Raman scattering (dual-CARS) microscopy. This new technique permits simultaneous imaging of two species characterized by different molecular vibrations, as well as the removal of nonresonant background. This is achieved by using three synchronized laser pulses probing two different vibrations. We demonstrate the virtues of the method by imaging a mixture of nondeuterated and deuterated lipids, clearly distinguishing the individual components and their organization in the mixed arrangement. Further, dual-CARS images of lipid stores in living Caenorhabditis elegans nematodes show that the suppression of the nonresonant background results in significantly enhanced image contrast.

Equipment Design↗

PubMed2004

Minimizing urine autofluorescence under multi-photon excitation conditions.

We report on the effects of excitation wavelength, laser power, and phase resolution on the multi-photon-excited autofluorescence (background) from human urine. When compared to the autofluorescence under one-photon excitation conditions (lambda(ex) = 260-480 nm), the urine multi-photon-excited autofluorescence (lambda(ex) = 725-950 nm) can be less complicated. However, at higher laser powers, the multi-photon-excited autofluorescence spectra that are produced by excitation above ~775 nm are more complex in comparison to the corresponding one-photon-excited autofluorescence. The origin of these more complex spectra arises from simultaneous two- and three-photon-driven excitation of intrinsic luminescent species within the urine. At lower laser powers, three-photon-driven processes are minimized and the autofluorescence spectrum is simplified. Phase resolution is used to further minimize the urine autofluorescence, but it cannot fully eliminate autofluorescence even when excitation is performed under multi-photon conditions at 950 nm. For detecting 250 nM Rhodamine 6G (a mock analyte) dissolved in urine, we find that the two-photon excitation is superior in comparison to one-photon excitation by 5- to 70-fold, depending on the excitation wavelength. Phase resolution combined with two-photon excitation leads to an additional 5- to 7-fold improvement in signal-to-background ratios in comparison to steady-state two-photon excitation.

Algorithms↗