Simultaneous spatial and temporal focusing of femtosecond pulses (original) (raw)
Related papers
Improved depth resolution in video-rate line-scanning multiphoton microscopy using temporal focusing
Optics Letters, 2005
In this study, a microscope based on spatiotemporal focusing offering widefield multiphoton excitation has been developed to provide fast optical sectioning images. Key features of this microscope are the integrations of a 10 kHz repetition rate ultrafast amplifier featuring high instantaneous peak power (maximum 400 J/pulse at a 90 fs pulse width) and a TE-cooled, ultra-sensitive photon detecting, electron multiplying charge-coupled camera into a spatiotemporal focusing microscope. This configuration can produce multiphoton images with an excitation area larger than 200 × 100 m 2 at a frame rate greater than 100 Hz (current maximum of 200 Hz). Brownian motions of fluorescent microbeads as small as 0.5 m were observed in real-time with a lateral spatial resolution of less than 0.5 m and an axial resolution of approximately 3.5 m. Furthermore, second harmonic images of chicken tendons demonstrate that the developed widefield multiphoton microscope can provide high resolution z-sectioning for bioimaging.
Extended depth of focus multiphoton microscopy via incoherent pulse splitting
We present a phase mask that can be easily added to any multi-photon raster scanning microscope to extend the depth of focus five-fold at a small loss in lateral resolution. The method is designed for ultrafast laser pulses or other light-sources featuring a low coherence length. In contrast to other methods of focus extension, our approach uniquely combines low complexity, high light-throughput and multicolor capability. We characterize the point-spread function in a two-photon microscope and demonstrate fluorescence imaging of GFP labeled neurons in fixed brain samples as imaged with conventional and extended depth of focus two-photon microscopy.
Chemical Physics Letters, 2016
Selective excitation of a particular fluorophore in an ensemble of different fluorophores with overlapping fluorescence spectra is shown to be dependent on the time delay of femtosecond pulse pairs in multiphoton fluorescence microscopy. In particular, the two-photon fluorescence behavior of the Texas Red and DAPI dye pair inside Bovine Pulmonary Artery Endothelial (BPAE) cells depends strongly on the center wavelength of the laser, as well as the delay between two identical laser pulses in one-color femtosecond pulse-pair excitation scheme. Thus, we present a novel design concept using pairs of femtosecond pulses at different central wavelengths and tunable pulse separations for controlling the image contrast between two spatially and spectrally overlapping fluorophores. This femtosecond pulse-pair technique is unique in utilizing the variation of dye dynamics inside biological cells as a contrast mode in microscopy of different fluorophores.
Mapping femtosecond pulse front distortion and group velocity dispersion in multiphoton microscopy
2006
Group velocity dispersion (GVD) and pulse front distortion of ultrashort pulses are of critical importance in efficient multiphoton excitation microscopy. Since measurement of the pulse front distortion due to a lens is not trivial we have developed an imaging interferometric cross-correlator which allows us to measure temporal delays and pulse-widths across the spatial profile of the beam. The instrument consists of a modified Michelson interferometer with a reference arm containing a voice-coil delay stage and an arm which contains the optics under test. The pulse replicas are recombined and incident on a 22×22 lenslet array. The beamlets are focused in a 0.5 mm thick BBO crystal (cut for Type I second harmonic generation), filtered to remove the IR component of the beam and imaged using a 500 fps camera. The GVD and pulse front distortion are extracted from the temporal stack of beamlet images to produce a low resolution spatio-temporal map.
Scanningless Depth Resolved Microscopy by Temporal Focusing of Ultrashort Pulses
1990
The ability to perform optical sectioning is one of the great advantages of laser- scanning microscopy, be it confocal or multiphoton microscopy. This comes however, at a cost of long image acquisition times, of an order of tens of milliseconds, due to the serial acquisition of data points. We show that by introducing spatiotemporal pulse shaping techniques it is possible to obtain full-frame depth resolved imaging without scanning, in a very simple setup. In multiphoton laser scanning microscopes, the depth resolution is achieved by spatially focusing an ultrashort pulse to achieve a high intensity at the focal plane (1). Due to the nonlinear dependence of the signal on the intensity this results in superb rejection of the out-of-focus signal. In contrast, our method relies on temporal focusing of the illumination pulse. The pulsed excitation field is compressed as it propagates through the sample, reaching its shortest duration (and highest peak intensity) at the focal plane, befo...
Microscopy with femtosecond laser pulses: applications in engineering, physics and biomedicine
Applied Surface Science, 2003
The combination of microscopy and femtosecond laser illumination turns out to be very attractive and useful for imaging in engineering, physics and biomedicine. The high laser intensity and low average power allow for the generation of nonlinear imaging signals that contain information complementary to classical imaging modes. The current state-of-the-art is reviewed and nonlinear current imaging and imaging of ballistic electron transport in Au-®lms is discussed in detail.
Selective two-photon microscopy with shaped femtosecond pulses
Optics Express, 2003
Selective two-photon excitation of fluorescent probe molecules using phase-only modulated ultrashort 15-fs laser pulses is demonstrated. The spectral phase required to achieve the maximum contrast in the excitation of different probe molecules or identical probe molecules in different micro-chemical environments is designed according to the principles of multiphoton intrapulse interference (MII). The MII method modulates the probabilities with which specific spectral components in the excitation pulse contribute to the two-photon absorption process due to the dependence of the absorption on the power spectrum of E 2 (t) [1]. Images obtained from a number of samples using the multiphoton microscope are presented.
Multiphoton microscopy system with a compact fiber-based femtosecond-pulse laser and handheld probe
Journal of biophotonics, 2011
We report on the development of a compact multiphoton microscopy (MPM) system that integrates a compact and robust fiber laser with a miniature probe. The all normal dispersion fiber femtosecond laser has a central wavelength of 1.06 μm, pulse width of 125 fs and average power of more than 1 W. A double cladding photonic crystal fiber was used to deliver the excitation beam and to collect the two-photon signal. The hand-held probe included galvanometer-based mirror scanners, relay lenses and a focusing lens. The packaged probe had a diameter of 16 mm. Second harmonic generation (SHG) images and two-photon excited fluorescence (TPEF) images of biological tissues were demonstrated using the system.
Widefield multiphoton and temporally decorrelated multifocal multiphoton microscopy
Optics Express, 2000
We demonstrate a widefield multiphoton microscope and a temporally decorrelated, multifocal, multiphoton microscope that is based on a high-efficiency array of cascaded beamsplitters. Because these microscopes use ultrashort pulse excitation over large areas of the sample, they allow efficient use of the high-average power available from modern ultrashort pulse lasers.