Endoscopic optical coherence tomography based on a microelectromechanical mirror

An endoscopic optical coherence tomography (OCT) system based on a microelectromechanical mirror to facilitate lateral light scanning is described. The front-view OCT scope, adapted to the instrument channel of a commercial endoscopic sheath, allows real-time cross-sectional imaging of living biological tissue via direct endoscopic visual guidance. The transverse and axial resolutions of the OCT scope are roughly 20 and 10.2mum, respectively. Cross-sectional images of 500x1000 pixels covering an area of 2.9 mmx2.8 mm can be acquired at ~5 frames/s and with nearly 100-dB dynamic range. Applications in thickness measurement and bladder tissue imaging are demonstrated.     

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.     

Miniaturized probe based on a microelectromechanical system mirror for multiphoton microscopy

A factor that limits the use of multiphoton microscopy (MPM) in clinical and preclinical studies is the lack of a compact and flexible probe. We report on a miniaturized MPM probe employing a microelectromechanical system (MEMS) scanning mirror and a double-clad photonic crystal fiber (DCPCF). The use of a MEMS mirror and a DCPCF provides many advantages, such as size reduction, rapid and precise scanning, efficient delivery of short pulses, and high collection efficiency of fluorescent signals. The completed probe was 1 cm in outer diameter and 14 cm in length. The developed probe was integrated into an MPM system and used to image fluorescent beads, paper, and biological specimens.     

基于同步辐射的快速扫描X射线微束荧光成像方法

在上海光源硬X射线微聚焦光束线站(BL15U1)上, 基于EPICS软件平台, 集成运动控制, 光强探测, 荧光探测等功能, 实现了"飞行"模式 (on-the-fly) X射线扫描微束荧光成像方法. 用"飞行"扫描X射线荧光成像法获得了标准镍网, 以及微量元素Cu, Zn,K, Fe在样品老鼠脾内的分布图像, 结果显示该方法不但在速度上有了极大的提高, 而且获得的元素分布图像具有高质量. 关键词： 快速扫描X射线微束荧光成像 同步辐射 微量元素分布     

Compact two-photon fluorescence microscope based on a single-mode fiber coupler

We present a two-photon fluorescence microscope based on a three-port single-mode optical fiber coupler. It is found that the coupler behaves as a low-pass filter that can deliver an ultrashort-pulsed laser beam of as much as 150 mW of power in the wavelength range from 770 to 870 nm as well as collect a two-photon fluorescence signal in the visible range. As a result of using the fiber coupler, the new two-photon imaging system exhibts a number of advantages, including a compact arrangement, freedom from vibration from lasers and electronic devices, self-alignment, reduction of multiple scattering, and an enhanced optical sectioning effect. The effectiveness of the new instrument is demonstrated with a set of three-dimensional images of biological samples. This instrument may make two-photon fluorescence endoscopy possible for in vivo medical applications.     

Simultaneous imaging of multiple focal planes using a two-photon scanning microscope

Despite all the advances in nonlinear microscopy, all existing instruments are constrained to obtain images of one focal plane at a time. In this Letter we demonstrate a two-photon absorption fluorescence scanning microscope capable of imaging two focal planes simultaneously. This is accomplished by temporally demultiplexing the signal coming from two focal volumes at different sample depths. The scheme can be extended to three or more focal planes.     

Zebrafish imaging and two-photon fluorescence imaging using ZnSe quantum dots

This study is to report a ZnSe quantum dot with a large two-photon absorption cross section and good biocompatibility,which can be used in bioimaging.Fluorescence emission at 410 nm is observed in the quantum dot under 760-nm laser excitation.These biocompatible quantum dots exhibit a two-photon cross-section of 9.1×10~5 GM(1 GM=10~(-50) cm~4·s/photon).Two-photon excited laser scanning microscopic images show that cells co-cultured with ZnSe quantum dots are found in the blue channel at a fluorescence intensity that is 14.5 times that of control cells not cocultured with quantum dots.After incubating zebrafish larvae with ZnSe quantum dots for 24 h,the fluorescence intensity of the yolk sac stimulated by ultraviolet light is 2.9 times that of the control group.The proposed material shows a great potential application in biological imaging.     

Flow imaging by use of femtosecond-laser-induced two-photon fluorescence

A novel technique is demonstrated for the imaging of turbulent flows in which a single window to the flow is the only optical access required. A femtosecond laser is used to excite two-photon fluorescence in a disodium-fluorescein-seeded water jet. The fluorescence signal is generated at only the focal point of the laser because of the highly nonlinear nature of the two-photon absorption, and it is collected in a direction counterpropagating to the excitation beam. Tight focusing of the laser is used to limit the probe volume, and the two-dimensional mean and rms concentration images are collected by raster scanning the laser.     

Combined Raman and continuous-wave-excited two-photon fluorescence cell imaging

We demonstrate a confocal optical microscope that combines cw two-photon-excited fluorescence microscopy with confocal Raman microscopy. With this microscope fast image acquisition with fluorescence imaging can be used to select areas of interest for subsequent chemical analysis with spontaneous Raman imaging. The distribution of the UV-absorbing fluorophore Hoechst 33342 in the apoptotic HeLa cells is measured in the combined cw two-photon-excited fluorescence and Raman microscopy modes. The 647-nm line of a Kr-ion laser is used to excite both the Raman scattering and the two-photon-excited fluorescence emission. The lateral and axial resolutions in the two imaging modes are compared by use of the Gaussian beam approximation and backprojection of the focal volume through the confocal pinhole.     

Biosensing based on two-photon fluorescence measurements through optical fibers

We have performed two-photon fluorescence detection in a new scheme in which femtosecond laser pulses were delivered thorugh an optical fiber for nonlinear excitation and the emitted fluorescence was collected through the same fiber. Single-mode fibers were determined to give higher detection efficiency than multimode fibers, consistent with theoretical considerations. The utility of fiber-optic sensing based on two-photon fluorescence detection was proved by an experiment that measured the uptake of a targeted drug delivery agent into cultured cancer cells.     

Photolytic-interference-free, femtosecond two-photon fluorescence imaging of atomic hydrogen

We discuss photolytic-interference-free, high-repetition-rate imaging of reaction intermediates in flames and plasmas using femtosecond (fs) multiphoton excitation. The high peak power of fs pulses enables efficient nonlinear excitation, while the low energy nearly eliminates interfering single-photon photodissociation processes. We demonstrate proof-of-principle, interference-free, two-photon laser-induced fluorescence line imaging of atomic hydrogen in hydrocarbon flames and discuss the method's implications for certain other atomic and molecular species.     

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.     

Bimorph deformable mirror based adaptive optics scanning laser ophthalmoscope for retina imaging in vivo

A bimorph deformable mirror(DM) with a large stroke of more than 30 μm using 35 actuators is presented and characterized for an adaptive optics(AO) confocal scanning laser ophthalmoscope application. Facilitated with a Shack–Hartmann wavefront sensor, the bimorph DM-based AO operates closed-loop AO corrections for human eyes and reduces wavefront aberrations in most eyes to below 0.1 μm rms. Results from living eyes, including one exhibiting ～5D of myopia and ～2D of astigmatism along with notable high-order aberrations, reveal a practical efficient aberration correction and demonstrate a great benefit for retina imaging, including improving resolution, increasing brightness, and enhancing the contrast of images.     

Experimental demonstration of phase correction with a 32 x 32 microelectromechanical systems mirror and a spatially filtered wavefront sensor

A 32 x 32 microelectromechanical systems deformable mirror is controlled in closed loop with a spatially filtered Shack-Hartmann wavefront sensor and a Fourier-transform wavefront reconstruction algorithm. A phase plate based on atmospheric turbulence statistics is used to generate a 1 microm peak-valley static phase aberration. Far-field images and direct phase measurements of the residual are used to compare performance with and without the spatial filter. Use of the spatial filter reduces error in the controllable band from 20 to 6 nm rms. Residual phase power is reduced by more than a factor of 5 for all spatial frequencies up to 0.85 x 1/2d, with a maximum attenuation factor of 37.     

Three-dimensional chemical concentration maps in a microfluidic device using two-photon absorption fluorescence imaging

Two-photon absorption fluorescence is employed within a microfluidic device to create a three-dimensional chemical concentration map for mixing uniformity characterization. This multiphoton technique images fluorescence intensity directly and provides a simple, rapid, and readily employed route to composition characterization within microfluidic systems.     

Aberration-correction results from a segmented microelectromechanical deformable mirror and a refractive lenslet array

Quadratic aberration is successfully corrected with a segmented microelectromechanical deformable mirror in conjunction with a refractive lenslet array. Use of the lenslet array greatly improves the effective fill factor of the correcting element. Experimental results show correction approaching the diffraction limit for an extreme spherical aberration.     

Quantitative imaging of mixing dynamics in microfluidic droplets using two-photon fluorescence lifetime imaging

Droplet-based microfluidic systems enable miniaturization of chemical reactions in femtoliter to picoliter volume compartments. Quantifying mixing dynamics of the reagents in droplets is critical to determine the system performance. In this Letter, we developed a two-photon excitation fluorescence lifetime imaging technique to quantitatively image the mixing dynamics in micro?uidic droplets. A cross/autocorrelation method was used to reconstruct a high-quality fluorescence lifetime image of the droplet. The fluorescence decay was analyzed for accurate determination of the mixing ratio at each pixel of the image.     

Use of a microelectromechanical mirror for adaptive optics in the human eye

Ophthalmic instrumentation equipped with adaptive optics offers the possibility of rapid and automated correction of the eye's optics for improving vision and for improving images of the retina. One factor that limits the widespread implementation of adaptive optics is the cost of the wave-front corrector, such as a deformable mirror. In addition, the large apertures of these elements require high pupil magnification, and hence the systems tend to be physically large. We present what are believed to be the first closed-loop results when a compact, low-cost, surface micromachined, microelectromechanical mirror is used in a vision adaptive-optics system. The correction performance of the mirror is shown to be comparable to that of a Xinetics mirror for a 4.6-mm pupil size. Furthermore, for a pupil diameter of 6.0-mm, the residual rms error is reduced from 0.36 to 0.12 microm and individual photoreceptors are resolved at a pupil eccentricity of 1 degrees from the fovea.     

Lateral scanning confocal microscopy for the determination of in-plane displacements of microelectromechanical systems devices

A method named lateral scanning confocal microscopy (LtSCM) is proposed with the aim of determining the in-plane displacement of microstructures, especially of those moving components within microelectromechanical systems (MEMS) actuators and sensors, which feature high aspect ratio and limited geometrical size in one or two dimensions within the surface plane. The principle of the LtSCM is presented and theoretically analyzed, which indicates that the LtSCM has the potential to determine with high resolution the in-plane displacement, position, and even the geometrical size of the object. Furthermore, in the case of in-plane displacement measurement, the measurement resolution of the LtSCM should be insensitive to the dynamic performance of the movable microstructures, i.e., from quasi-static to ultrahigh speed. In a proof-of-principle experiment, the voltage-displacement response of an electrostatic comb-drive actuator has been obtained with nanometric resolution.