Simultaneous imaging of human cone mosaic with adaptive optics enhanced scanning laser ophthalmoscopy and high-speed transversal scanning optical coherence tomography

We describe a novel instrument capable of acquiring, simultaneously, adaptive optics enhanced scanning laser ophthalmoscopy and optical coherence tomography (OCT) images of the human cone mosaic in vivo. The OCT system is based on transversal scanning of the sample with a line scan rate of 14 kHz, approximately 20 times faster than a previously reported instrument. We demonstrate the capability of this instrument with the measurement of the human cone spacing in perifoveal retina.     

Directional imaging of the retinal cone mosaic

We describe a near-IR scanning laser ophthalmoscope that allows the retinal cone mosaic to be imaged in the human eye in vivo without the use of wave-front correction techniques. The method takes advantage of the highly directional quality of cone photoreceptors that permits efficient coupling of light to individual cones and subsequent detection of most directional components of the backscattered light produced by the light-guiding effect of the cones. We discuss details of the system and describe cone-mosaic images obtained under different conditions.     

Artifact removal in Fourier-domain optical coherence tomography with a piezoelectric fiber stretcher

We describe an artifact removal setup swept-source optical coherence tomography (OCT) system that enables high-speed full-range imaging. We implement a piezoelectric fiber stretcher to generate a periodic phase shift between successive A-scans, thus introducing a transverse modulation. The depth ambiguity is then resolved by performing a Fourier filtering in the transverse direction before processing the data in the axial direction. The dc artifact is also removed. The key factor is that the piezoelectric fiber stretcher can be used to generate discrete phase shifts with a high repetition rate. The proposed experimental setup is a much improved version of the previously reported B-M mode scanning for spectral-domain OCT in that it does not generate additional artifacts. It is a simple and low-cost solution for artifact removal that can easily be applied.     

Phase-resolved optical coherence tomography and optical Doppler tomography for imaging blood flow in human skin with fast scanning speed and high velocity sensitivity

We have developed a novel phase-resolved optical coherence tomography (OCT) and optical Doppler tomography (ODT) system that uses phase information derived from a Hilbert transformation to image blood flow in human skin with fast scanning speed and high velocity sensitivity. Using the phase change between sequential scans to construct flow-velocity imaging, this technique decouples spatial resolution and velocity sensitivity in flow images and increases imaging speed by more than 2 orders of magnitude without compromising spatial resolution or velocity sensitivity. The minimum flow velocity that can be detected with an axial-line scanning speed of 400 Hz and an average phase change over eight sequential scans is as low as 10 microm/s, while a spatial resolution of 10 microm is maintained. Using this technique, we present what are to our knowledge the first phase-resolved OCT/ODT images of blood flow in human skin.     

Adaptive-optics ultrahigh-resolution optical coherence tomography

Merging of ultrahigh-resolution optical coherence tomography (UHR OCT) and adaptive optics (AO), resulting in high axial (3 microm) and improved transverse resolution (5-10 microm) is demonstrated for the first time to our knowledge in in vivo retinal imaging. A compact (300 mm x 300 mm) closed-loop AO system, based on a real-time Hartmann-Shack wave-front sensor operating at 30 Hz and a 37-actuator membrane deformable mirror, is interfaced to an UHR OCT system, based on a commercial OCT instrument, employing a compact Ti:sapphire laser with 130-nm bandwidth. Closed-loop correction of both ocular and system aberrations results in a residual uncorrected wave-front rms of 0.1 microm for a 3.68-mm pupil diameter. When this level of correction is achieved, OCT images are obtained under a static mirror configuration. By use of AO, an improvement of the transverse resolution of two to three times, compared with UHR OCT systems used so far, is obtained. A significant signal-to-noise ratio improvement of up to 9 dB in corrected compared with uncorrected OCT tomograms is also achieved.     

Integrated multimodal endomicroscopy platform for simultaneous en face optical coherence and two-photon fluorescence imaging

We report an all-fiber-optic scanning, multimodal endomicroscope capable of simultaneous optical coherence tomography (OCT) and two-photon fluorescence (TPF) imaging. Both imaging modalities share the same miniature fiber-optic scanning endomicroscope, which consists of a double-clad fiber with a core operating in single mode at both the OCT (1310 nm) and two-photon excitation (1550 nm) wavelengths, a piezoelectric two-dimensional fiber-optic beam scanner, and a miniature aspherical compound lens suitable for simultaneous acquisition of en face OCT and TPF images. A fiber-optic wavelength division multiplexer was employed in the integrated platform to combine the low coherence OCT light source and the femtosecond two-photon excitation laser into the same optical path. Preliminary imaging results of cell cultures and mouse tissue ex vivo demonstrate the feasibility of simultaneous real-time OCT and TPF imaging in a scanning endomicroscopy setting for the first time.     

Imaging of subcutaneous blood vessels and flow velocity profiles by optical coherence tomography

We have applied a compact low power rapid scanning Doppler Optical Coherence Tomography system to monitor multi-dimensional velocity profiles within the complex vessels and simultaneous real-time non-invasive imaging of skin tissues morphology in vivo, in the wavelength range of 1.3–1.5 nm. Optical clearing of skin tissues has been utilized to achieve depth of OCT images up to 1.7 mm. Current approach enables applying low-power (0.4–0.5 mW) and low-noise broadband near-infrared light sources and obtaining OCT images with down to 12 μm spatial resolution. Two-dimensional time-domain OCT images of complex flow velocity profiles in blood vessel phantom and in vivo subcutaneous human skin tissues are presented. The effect of optical clearing on in vivo images is demonstrated and discussed.     

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.     

Optically deviated focusing method based high-speed SD-OCT for in vivo retinal clinical applications

The aim of this study is to provide accurately focused, high-resolution in vivo human retinal depth images using an optically deviated focusing method with spectral-domain optical coherence tomography (SD-OCT) system. The proposed method was applied to increase the retinal diagnosing speed of patients with various values of retinal distances (i.e., the distance between the crystalline eye lens and the retina). The increased diagnosing speed was facilitated through an optical modification in the OCT sample arm configuration. Moreover, the optical path length matching process was compensated using the proposed optically deviated focusing method. The developed system was mounted on a bench-top cradle to overcome the motion artifacts. Further, we demonstrated the capability of the system by carrying out in vivo retinal imaging experiments. The clinical trials confirmed that the system was effective in diagnosing normal and abnormal retinal layers as several retinal abnormalities were identified using non-averaged single-shot OCT images, which demonstrate the feasibility of the method for clinical applications.     

Intraoperative spectral domain optical coherence tomography for vitreoretinal surgery

We demonstrate in vivo human retinal imaging using an intraoperative microscope-mounted optical coherence tomography system (MMOCT). Our optomechanical design adapts an Oculus Binocular Indirect Ophthalmo Microscope (BIOM3), suspended from a Leica ophthalmic surgical microscope, with spectral domain optical coherence tomography (SD-OCT) scanning and relay optics. The MMOCT enables wide-field noncontact real-time cross-sectional imaging of retinal structure, allowing for SD-OCT augmented intrasurgical microscopy for intraocular visualization. We experimentally quantify the axial and lateral resolution of the MMOCT and demonstrate fundus imaging at a 20Hz frame rate.     

Homodyne en face optical coherence tomography

We demonstrate, for what we believe to be the first time, the use of a 3 x 3 fiber-optic coupler to realize a homodyne optical coherence tomography (OCT) system for en face imaging of highly scattering tissues and turbid media. The homodyne OCT setup exploits the inherent phase shifts between different output ports of a 3 x 3 fiber-optic coupler to extract amplitude information of a sample. Our homodyne en face OCT system features a measured resolution of 14 microm axially and 9.4 microm laterally with a 90 dB signal-to-noise ratio at 10 micros integration time. En face OCT imaging of a stage 52 Xenopus laevis was successfully demonstrated at a depth of 600 microm within the sample.     

Molecular contrast in optical coherence tomography by use of a pump-probe technique

We describe a novel technique for contrast enhancement in optical coherence tomography (OCT) that makes possible molecular-specific imaging for what is believed to be the first time. A pump-probe technique is employed in which a pulsed pump laser is tuned to ground-state absorption in a molecule of interest. The location of the target molecule population is derived from the resulting transient absorption of OCT sample-arm light acting as probe light. A signal processing technique for three-dimensional localization of the transient absorption signal is described, and preliminary results exhibiting OCT contrast from methylene blue dye in multilayer and scattering phantoms are presented.     

Simultaneous optical coherence tomography--Indocyanine Green dye fluorescence imaging system for investigations of the eye's fundus

We have developed a dual-channel optical coherence tomography-Indocyanine Green dye (OCT-ICG) fluorescence system based on a previously reported ophthalmic OCT confocal imaging system. The confocal channel is tuned to the fluorescence wavelength range of the ICG, and light from the same optical source is used to generate the OCT image and to excite the ICG fluorescence. The system enables the clinician to visualize simultaneously en face OCT slices and corresponding ICG angiograms of the ocular fundus, displayed side by side. C-scan (constant depth) and B-scan (cross section) images are collected by a fast en face scan (T scan). The pixel-to-pixel correspondence between the OCT and angiography images allows the user to capture OCT B scans precisely at selected points on the ICG confocal images.     

Use of a scanner to modulate spatial interferograms for in vivo full-range Fourier-domain optical coherence tomography

We report a new yet simple method to achieve full-range complex Fourier-domain optical coherence tomography (OCT) for in vivo imaging. The method utilizes a scanner that is dedicated for lateral scanning in the system to introduce a constant carrier frequency into the OCT spectral interferograms during the scanning. This is achieved by simply offsetting the sampling beam spot away from the pivot point of the scanning mirror. We demonstrate the method experimentally for in vivo full-range imaging of the anterior segment of a human eye. The method is free from complex conjugate mirror image and self-cross-correlation image artifacts.     

Combined confocal/en face T-scan-based ultrahigh-resolution optical coherence tomography in vivo retinal imaging

Combined confocal scanning ophthalmoscopy/en face T-scan-based ultrahigh-resolution optical coherence tomography (OCT) of the human retina in vivo is reported for the first time to our knowledge. The system uses a superluminescent diode-based broadband source, which gives an axial resolution of 3.2 microm in the retina. We demonstrate acquisition of T-scan-based OCT B-scan and simultaneous confocal/C-scan images of the human retina of large lateral size (covering a field of up to 20 degrees ) at a frame rate of 2Hz.     

Simultaneous swept source optical coherence tomography of the anterior segment and retina using coherence revival

We report on an implementation of coherence revival-based heterodyne swept source optical coherence tomography that is capable of simultaneously imaging the anterior and posterior eye. A polarization-encoded sample arm was used to efficiently focus orthogonal polarizations on the anterior segment and retina. Depth encoding was achieved using coherence revival, which allows for multiple depths within a sample to be simultaneously imaged and frequency encoded by carefully controlling the optical pathlength of each sample path. This design is a significant step toward whole-eye optical coherence tomography (OCT), which would enable customized ray-traced modeling of patient eyes to improve refractive surgical interventions and eliminate optical artifacts in retinal OCT diagnostics. We demonstrated the feasibility of this system for in vivo imaging by simultaneously acquiring images of the anterior segments and retinas in healthy human volunteers.     

Three-dimensional non-destructive optical evaluation of laser-processing performance using optical coherence tomography

We demonstrate the use of optical coherence tomography (OCT) as a non-destructive diagnostic tool for evaluating laser-processing performance by imaging the features of a pit and a rim. A pit formed on a material at different laser-processing conditions is imaged using both a conventional scanning electron microscope (SEM) and OCT. Then using corresponding images, the geometrical characteristics of the pit are analyzed and compared. From the results, we could verify the feasibility and the potential of the application of OCT to the monitoring of the laser-processing performance.     

Tracking optical coherence tomography

An experimental tracking optical coherence tomography (OCT) system has been clinically tested. The prototype instrument uses a secondary sensing beam and steering mirrors to compensate for eye motion with a closed-loop bandwidth of 1 kHz and tracking accuracy, to within less than the OCT beam diameter. The retinal tracker improved image registration accuracy to <1 transverse pixel (<60 microm). Composite OCT images averaged over multiple scans and visits show a sharp fine structure limited only by transverse pixel size. As the resolution of clinical OCT systems improves, the capability to reproducibly map complex structures in the living eye at high resolution will lead to improved understanding of disease processes and improved sensitivity and specificity of diagnostic procedures.     

Fourier domain optical coherence tomography using optical demultiplexers imaging at 60,000,000 lines/s

We describe high-speed Fourier domain optical coherence tomography (OCT) using optical demultiplexers (ODs) for spectral dispersion. The OD enables separation of a narrow spectral band of 14 GHz (0.11 nm) from a broadband incident light at 256 different frequencies in 25.0 GHz intervals centered at 192.2 THz (1559.8 nm). OCT imaging of 60,000,000 axial scans per second was achieved through parallel signal acquisition using 256 balanced photoreceivers to simultaneously detect all the output signals from the ODs in a Fourier domain OCT system. OCT imaging at a 16 kHz frame rate, 1100 A-lines per frame, 3 mm depth range, and 23 microm resolution was demonstrated using a resonant scanner for lateral scanning.     

Statistics and reduction of speckle in optical coherence tomography

Studies have shown that optical coherence tomography (OCT) is useful in imaging microscopic structures through highly scattering media. Because spatially coherent light is used in OCT, speckle in the reconstructed image is unavoidable, resulting in degradation of the quality of the OCT images and impaired ability to differentiate subsurface structures. Therefore speckle reduction is an important issue in OCT imaging. We develop speckle statistics that are appropriate to the OCT measurements and demonstrate a simple and practical speckle-reduction technique.