Intravascular polarization-sensitive optical coherence tomography (IV-PSOCT) provides depth-resolved tissue birefringence which can be used to evaluate the mechanical stability of a plaque. Here, we demonstrate a very simple method of constructing an intravascular polarization-sensitive optical coherence tomography (IV-PSOCT) system. A conventional intravascular OCT system can be modified for IV-PSOCT by applying a 12-m polarization-maintaining fiber based imaging probe. Its two polarization modes separately produce OCT images of polarization detection channels spatially distinguished by an image separation of 2.7 mm. We experimentally validated our IV-PSOCT with chicken tendon, chicken breast, and coronary artery for the image samples. We found that the birefringent properties can be successfully visualized by our endoscopic imaging tool.
KEYWORDS: Single photon, Photons, Time resolved spectroscopy, Time metrology, Temporal resolution, Photomultipliers, Electronics, Digital photography, Digital electronics
Phase stability of an optical coherence elastography (OCE) system is the key determining factor for precision elasticity measurement. In this study, we developed an OCE system based on swept-source optical coherence tomography (SS-OCT) with a common-path configuration (SS-OCECP). Our system has a phase stability of 4.2 mrad without external stabilization or extensive post-processing, such as averaging. This phase stability allows us to detect a displacement as small as ~300 pm. We validated the SS-OCECP performance in a tissue-mimicking phantom and an in vivo rabbit model, and the results demonstrated significant improved phase stability compared to conventional SS-OCE. To the best of our knowledge, we demonstrated the first SS-OCECP system, which possess high phase stability and can be utilized to significantly improve the sensitivity of elastography.
Degenerative joint disease (DJD) is a disease that the articular cartilage changes from hyaline cartilage to fibrous cartilage. PS-OCT may provide a method to quantitatively analyze cartilage birefringence and diagnose DJD. Here, we proposed a novel PS-OCT system that uses spun fiber to construct the sample arm. In this work, phase retardation map, optical axis map, and conventional OCT images of hyaline cartilage and fibrous cartilage are presented, and the differences in the birefringence of these two types of cartilage are identified. The proposed PS-OCT system demonstrates great potential for accurate diagnosis of DJD in the clinic.
Polarization sensitive optical coherence tomography (PS-OCT) is a functional extension of optical coherence tomography (OCT). It provides addition information of the sample based on by analyzing polarization states of the backscattering light. Serval PS-OCT such as free-space optics, single model fiber and polarization maintaining (PM) fibers systems have been developed so far. However, the free space PS-OCT is prone to systematic errors and impractical in clinic settings. Dues to the uncontrolled polarization states, single model-based optics requires additional compensation and self-calibration techniques are required. Traditional PM fiber preserves the polarization states but found expensive and can not be used in rotating endoscopic or catheter probe. In this study, we develop a novel scheme of PS-OCT implementation using specific PM fiber, known as spun fiber, which has a structure of PM fiber twisted along the fiber optic axis and distinguishes two circular opalization states with different propagation speeds. Spun fiber has the advantages in maintaining the polarization states and regardless of fiber bends. The orientation insensitivity of the spun fiber makes it of great potential in endoscopic PS-OCT system. We tested our spun fiber-based PS-OCT system on chicken breast sample. The phase retardation image shows clear muscle structures compared to intensity-based OCT images, indicating our PS-OCT system has the ability to detect tissue birefringence.
KEYWORDS: Fluorescence lifetime imaging, Sensors, Luminescence, Photodetectors, Signal detection, Monte Carlo methods, Single photon, Analog electronics, Physics, Temporal resolution
GaAsP hybrid detectors, which is new kind of photodetector, has been known as its excellent performance in time correlated single photon counting technique. We have verified that this detector also shows excellent performance in analog mean-delay method, which is another kind of time-domain FLIM, so one can expect enhancement of performance in time-domain FLIM when using the hybrid detector.
Phasor plot analysis is one of the most powerful analysis technique in fluorescence lifetime imaging microscopy, especially for analysis of heterogeneous mixtures. Compared to frequency domain fluorescence lifetime measurement, time domain measurement offers information in various frequencies at once measurement, but needs high frequency sampling for stable signal acquisition, which requires a lot of memory in hardware and a long time for analysis, furthermore in TCSPC, acquisition time is extremely long due to low photon count rate. We suggest a new system with low pass filter, which leads to about 100 times faster measurement speed while maintaining precision and accuracy in usual modulation frequency.
KEYWORDS: Luminescence, Fluorescence lifetime imaging, Analog electronics, Signal processing, Signal detection, Super resolution, Photodetectors, Convolution, Microscopy, Fluorescence resonance energy transfer
Fluorescence lifetime imaging microscopy (FLIM) is a powerful imaging tool widely used in monitoring cells, organelles, and tissues in biosciences. Since fluorescence lifetimes of most probes are a few nanoseconds, 20 ps measurement resolution is normally required. This requirement is quite challenging even with the fastest available optical and electronic devices, and several brilliant time-domain super-resolution techniques have been proposed for FLIM. The analog mean-delay (AMD) method is a recently introduced time-domain super-resolution technique for FLIM. Detailed constraints in the AMD method and their impact on the performance of the AMD super-resolution lifetime measurement are presented with experiments and simulations.
The Analog mean delay (AMD) method is a multiphoton detection fluorescence lifetime measurement method. We have investigated the effect of the linearity of a photodetector response on the performance of the AMD method, which is a multiphoton detected fluorescence lifetime measurement method. A Monte-Carlo simulation scheme was adapted to generate various electronic signals for the AMD method with different linearity conditions of a photodetector. It is found that the photon economy is better if a photodetector with better linearity is used especially when the number of detected photons per pulse is low.
Deep anterior lamellar keratoplasty (DALK) is an emerging surgical technique for the restoration of corneal clarity and vision acuity. The big-bubble technique in DALK surgery is the most essential procedure that includes the air injection through a thin syringe needle to separate the dysfunctional region of the cornea. Even though DALK is a well-known transplant method, it is still challenged to manipulate the needle inside the cornea under the surgical microscope, which varies its surgical yield. Here, we introduce the DALK protocol based on the position-guided needle and M-mode optical coherence tomography (OCT). Depth-resolved 26-gage needle was specially designed, fabricated by the stepwise transitional core fiber, and integrated with the swept source OCT system. Since our device is feasible to provide both the position information inside the cornea as well as air injection, it enables the accurate management of bubble formation during DALK. Our results show that real-time feedback of needle end position was intuitionally visualized and fast enough to adjust the location of the needle. Through our research, we realized that position-guided needle combined with M-mode OCT is a very efficient and promising surgical tool, which also to enhance the accuracy and stability of DALK.
We present a new low-nonlinearity fiber of mode-filtered large-core fiber for flexible beam delivery of intense pulsed
light aiming at multi-photon endoscopy application. A multimode fiber of a large core diameter (20 μm) equips a mode
filtering means in the middle of the fiber link to suppress the high-order modes selectively. A large effective core area of
~200 μm2 has been achieved at 0.8-μm and 1.0-μm bands. This is 8 times larger than the core area of a conventional
SMF used for those spectral bands. Various advantages of our large-mode area fiber will be demonstrated and discussed
in this report.
Multi point fluorescence measurement system using basic image shifting method and commercial multi mode fiber is
presented in this paper. Using a singlet lens, the original fluorescence image of a sample is shifted to another plane which
can be monitored using ccd, and at the first image plane independent two fiber tips in an xyz stage deliver each
fluorescent signal at a specific sample position to a fluorescence correlation spectroscopy (FCS) with an electron
multiplying charge coupled device (EMCCD). The FCS is composed with an EMCCD, which can detect single molecule
level fluorescence light. Applying region of interest (ROI) and pixel binning, a time resolution of up to 2 ms can be
achieved, which is sufficient to resolve the diffusion of fluorescence micro-sphere in solution. The advantages of
implementing EMCCD cameras in wide-field ultra low light imaging, as well as in site-specific multi-point fluorescence
measurement system, can consequently also be exploited for spatially and spectrally resolved FCS. Experimental results
about FCS with spectrum informations demonstrate the advantage of the simplicity and flexibility of our system. We
expect that this multi point measurement system also can be applied to other study of bio molecular dynamics.
Confocal laser scanning microscopy (CLSM) has become the tool of choice for high-contrast fluorescence imaging in the
study of the three-dimensional and dynamic properties of biological system. However, the high cost and complexity of
commercial CLSMs urges many researchers to individually develop low cost and flexible confocal microscopy systems.
The high speed scanner is an influential factor in terms of cost and system complexity. Resonant galvo scanners at
several kHz have been commonly used in custom-built CLSMs. However, during the repeated illumination for live cell
imaging or 3D image formation, photobleaching and image distortion occurred at the edges of the scan field may be
more serious than the center due to an inherent property (e.g. sinusoidal angular velocity) of the scan mirror. Usually, no
data is acquired at the edges due to large image distortion but the excitation beam is still illuminated. Here, we present
the photobleaching property of CLSM with masked illumination, a simple and low cost method, to exclude the
unintended excitation illumination at the edges. The mask with a square hole in its center is disposed at the image plane
between the scan lens and the tube lens in order to decrease photobleaching and image distortion at the edges. The
excluded illumination section is used as the black level of the detected signals for a signal quantizing step. Finally, we
demonstrated the reduced photobleaching at the edges on a single layer of fluorescent beads and real-time image
acquisition without a standard composite video signal by using a frame grabber.
We present a new high-speed lifetime measurement scheme of analog mean-delay (AMD) method which is suitable for
studying dynamical time-resolved spectroscopy and high-speed fluorescence lifetime imaging microscopy (FLIM). In
our lifetime measurement method, the time-domain intensity of a decaying fluorescence light source is acquired as an
analog waveform, and the lifetime information of the source is extracted from the calculated mean temporal delay of the
waveform.
KEYWORDS: Luminescence, Photons, Analog electronics, Signal detection, Fluorescence lifetime imaging, Signal processing, Picosecond phenomena, Photodetectors, Data acquisition, Interference (communication)
We present a novel method for high-speed measurements of fluorescence lifetime, in which fluorescence signal for
precise lifetime determination is acquired in a short time on the order of microseconds. Our method is based on analog
signal that contains a number of fluorescence photons in a pulse, on the contrary to the conventional time-correlated
single-photon counting in which only a single photon is permitted for a fluorescence pulse. Because this method does not
have any problem of photon counting pile-up, the measurement speed is not limited by the single-photon constraint and
can increase up to the excitation repetition rate. In order to extract the accurate lifetime information from the analog
signal contaminated by the slow instrumental response function (IRF), we have developed a new signal processing
method, in which the lifetime is determined by difference between mean arrival time of the analog photo-electronic pulse
of fluorescence signal and one of IRF signal. By both experimental and theoretical studies, we have verified that the
measurement accuracy and precision are nearly independent of the width of the IRF so that inexpensive narrowbandwidth
photo-detectors and low-speed electronics can be used for this method. Excellent accuracy and precision have
been obtained experimentally for high-speed measurements completed in a few microseconds. These results suggest that
our method can be well applied to measurement of fast dynamic phenomena and real time fluorescence lifetime imaging
microscope with low cost.
We present a simple 2D image acquisition technique electronically implemented for laser scanning confocal microscope
using galvanometer scanners. In order to synchronize image acquisition process with the movement of the galvanometer
scanner, position signal of the mirror of the galvanometer scanner is used and manipulated for generating of sync-signals.
This is achieved using an analog-digital converter to read a position signal from the scanner which tells about its position
and to generate a trigger signal (or pixel clock) which tells the moment of digitizing the received analog signal from the
photo-detector. This facilitates processing the image in synchronization with the actual motion of the scanning laser
beam scanner. Image construction is performed by a video acquisition board (frame grabber). The newly developed
scanning and image acquisition systems are implemented in a confocal microscope with fiber-optic components for
compact configuration and flexible light path.
We present a fluorescence lifetime imaging microscope (FLIM) based on a real-time waveform acquisition method. The
fluorophores were excited by a 635-nm gain-switched laser diode, which produced short pulses with duration ~50 ps in a
20-MHz repetition rate. The fluorescence signals were detected by a silicon avalanche photo-diode (APD) in addition to
a wide-band electric amplifier. The converted electric pulses were sampled by a high-speed digitizer of which sampling
rate was 2 GS/s. In order to reduce the sampling interval for analyzing sub-nanosecond lifetimes, an interleaved data
acquisition technique was used. The effective sampling rate was increased to 10 GS/s. In addition, the impulse response
was measured simultaneously with the lifetime signals by an interleaving manner and was used in calibration of the
system. By using these methods, accurate lifetime information was acquired in a short time less than 8 μs.
We have demonstrated a Fourier-domain optical coherence tomography (FD-OCT) scheme with a high-speed
frequency-swept light source based on a chirped supercontinuum pulse. Instead of using a swept laser, an ultra-wideband
supercontinuum pulse was chirped or stretched in the time domain by using a long dispersive single-mode
optical fiber by the help of the group velocity dispersion. The chirped pulse was used directly as frequency-swept light
for OCT after measuring the relationship between the time delay and the wavelength. Very high acquisition speeds up to
5-MHz in A-line scan rate were achieved because there is no speed-limiting moving part in this scheme. And high
resolution up to 3.6 &mgr;m in air was enabled owing to the use of wideband supercontinuum. It was shown that the scheme
does not require re-calibration of the sweep characteristics because the sweeping mechanism is passive and stable.
We present an ultra-wideband supercontinuum source using a dispersion-shifted fiber and an amplified diode-laser pulse source. A gain-switched DFB laser operating at 1550-nm wavelength, which provides 30-ps pulses, was used for generating the seeding pulses. And serially cascaded low-cost EDFAs were employed to boost the peak power of the pulses to more than 1 kW. Single-mode supercontinuum spanning nearly the full near-IR band was obtained by passing the amplified pulses through a dispersion-shifted fiber. By investigating the characteristics of the generated supercontinuum pulses, the walk-off between the spectral components was found to limit the effective interaction length of the spectrum-broadening effects. In order to expand further the spectral range of the output, we have examined the time-gating ASE suppression scheme and use of a high-power EDFA. And the resulted outputs have reached wavelengths of 0.8 and 0.9 μm, respectively at the short-wavelength edges. Only the blue-shifted part that can be obtained using a short-wavelength-pass filter can exhibit 3-dB bandwidth more than 500 nm in the vicinity of 1.2 μm. The supercontinuum generation scheme provides a compact and reliable way to generate ultra-wideband flat spectrum that can be useful for high-resolution OCT.
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