Multispectral photoacoustic microscopy and optical coherence tomography using a single supercontinuum source.

We report on the use of a single supercontinuum (SC) source for multimodal imaging. The 2-octave bandwidth (475-2300 nm) makes the SC source suitable for optical coherence tomography (OCT) as well as for multispectral photoacoustic microscopy (MPAM). The IR band centered at 1310 nm is chosen for OCT to penetrate deeper into tissue with 8 mW average power on the sample. The 500-840 nm band is used for MPAM. The source has the ability to select the central wavelength as well as the spectral bandwidth. An energy of more than 35 nJ within a less than 50 nm bandwidth is achieved on the sample for wavelengths longer than 500 nm. In the present paper, we demonstrate the capabilities of such a multimodality imaging instrument based on a single optical source. In vitro mouse ear B-scan images are presented.

Speckle variance OCT for depth resolved assessment of the viability of bovine embryos.

The morphology of embryos produced by in vitro fertilization (IVF) is commonly used to estimate their viability. However, imaging by standard microscopy is subjective and unable to assess the embryo on a cellular scale after compaction. Optical coherence tomography is an imaging technique that can produce a depth-resolved profile of a sample and can be coupled with speckle variance (SV) to detect motion on a micron scale. In this study, day 7 post-IVF bovine embryos were observed either short-term (10 minutes) or long-term (over 18 hours) and analyzed by swept source OCT and SV to resolve their depth profile and characterize micron-scale movements potentially associated with viability. The percentage of en face images showing movement at any given time was calculated as a method to detect the vital status of the embryo. This method could be used to measure the levels of damage sustained by an embryo, for example after cryopreservation, in a rapid and non-invasive way.

Image quality improvement in optical coherence tomography using Lucy-Richardson deconvolution algorithm.

Optical coherence tomography (OCT) has the potential for skin tissue characterization due to its high axial and transverse resolution and its acceptable depth penetration. In practice, OCT cannot reach the theoretical resolutions due to imperfections of some of the components used. One way to improve the quality of the images is to estimate the point spread function (PSF) of the OCT system and deconvolve it from the output images. In this paper, we investigate the use of solid phantoms to estimate the PSF of the imaging system. We then utilize iterative Lucy-Richardson deconvolution algorithm to improve the quality of the images. The performance of the proposed algorithm is demonstrated on OCT images acquired from a variety of samples, such as epoxy-resin phantoms, fingertip skin and basaloid larynx and eyelid tissues.

Optical coherence tomography.

The review provides a concise explanation of principles of operation of different optical coherence tomography methods. A comparative analysis of their advantages and disadvantages is presented in relation to specific applications. The review will assist the reader in making an educated choice on the most suitable optical coherence tomography method to be used in a particular application.

Imaging of periocular basal cell carcinoma using en face optical coherence tomography: a pilot study.

AIM: To use en face optical coherence tomographic (OCT) imaging to identify features of tumour tissue and their correlation with histopathologic findings and to assess the effect of different wavelengths and resolutions of OCT in identifying tumour boundaries and features. METHODS: Excision specimens of consecutive biopsy-proven periocular basal cell carcinomas (BCCs) (n=8) were assessed by OCT, performing in vitro cross-section and en face scans of the tissues. Images were collected from three different machines: systems 1 and 2 had a wavelength of 1300 nm, and system 3 had a wavelength of 840 nm. System 2 used high numerical aperture interface optics that determines higher magnification and hence allows higher transversal resolution. All the eight specimens subsequently underwent routine histopathologic examination. RESULTS: Three common features of tumour tissue were observed in all the three systems: (1) lobular pattern of abnormal architecture, (2) dilated blood vessels and (3) high reflective margins. We compared the three systems based on their ability to pick up the three above-mentioned tumour features. In this respect, system 2 had the highest capability in picking up feature 1, followed by systems 1 and 3. In feature 2, similar results were obtained with all the three systems. System 3 was unable to pick up feature 3, whereas systems 1 and 2 performed equally. CONCLUSION: En face OCT imaging has the potential to identify tumour tissue from healthy tissue. It also showed correlation with corresponding histopathologic findings. Non-contact OCT imaging of the skin is a non-invasive and convenient method and can be useful for demarcating BCCs on the face and eyelids. Future larger studies on in vivo BCCs using en face ultra-high-resolution OCT should provide information on subtyping BCCs.

Combining SLO and OCT technology.

A review is presented of the research on high resolution imaging of the eye which can provide a dual display of images with different depth resolutions. The review refers to the flying spot scanning concept, widely exploited in the confocal scanning laser ophthalmoscope and recently extended to optical coherence tomography (OCT) imaging. For several reasons as presented in the paper, imaging with two different depth resolutions is useful and this has triggered the development of the dual en-face OCT--confocal imaging technology and of the OCT/Ophthalmoscope instrument. The dual acquisition and presentation can be performed either simultaneously (practised in the OCT/Ophthalmoscope) or sequentially. The sequential dual en-face OCT--confocal imaging technology can be implemented in different configurations and has specific applications. When the sequential switching is performed at the line rate of the raster frame, the display of the two images, OCT and confocal is quasi - simultaneous, in which case similar functionality is achieved to that of an OCT/Ophthalmoscope.

Optical coherence tomography.

A review is presented of different scanning, acquisition and processing techniques used to obtain depth-resolved information in optical-coherence tomography (OCT). The principles and performances of different OCT techniques are discussed and images from different types of tissue are presented. Special attention is devoted to the progress in using the time-domain flying spot OCT technique and combination of the en face OCT imaging with confocal microscopy. Although OCT is based on white light interferometry, which is a well established and an old technology, the quest for higher resolution and faster acquisition of in vivo images has ensured OCT a rapid evolution in the last decade. Highly adventurous avenues to expand the OCT capabilities and trends are presented at the end of the review.

En face optical coherence tomography: a new method to analyse structural changes of the optic nerve head in rat glaucoma.

AIM: To investigate en face optical coherence tomography (eOCT) and its use as an effective objective technique for assessing changes in the glaucomatous rat optic nerve head (ONH) in vivo, and compare it with confocal scanning laser ophthalmoscopy (cSLO). METHODS: 18 Dark Agouti (DA) rats with surgically induced ocular hypertension were imaged with eOCT and cSLO at regular intervals. Assessment included three dimensional (3D) topographic reconstructions, intensity z-profile plots, a new method of depth analysis to define a "multilayered" structure, and scleral canal measurements, in relation to the degree of intraocular pressure (IOP) exposure. RESULTS: The increased depth resolution of the eOCT compared to the cSLO was apparent in all methods of analysis, with better discrimination of tissue planes. This was validated histologically. eOCT demonstrated several significant changes in imaged rat ONH which correlated with IOP exposure, including the area of ONH (p<0.01), separation between retinal vessel and scleral layers (p<0.05), and anterior scleral canal opening expansion (p<0.05). CONCLUSION: eOCT appears to be effective in assessing rat ONH, allowing detailed structural analysis of the multilayered ONH structure. As far as the authors are aware, this is the first report of scleral canal expansion in a rat model. They suggest eOCT as a novel method for the detection of early changes in the ONH in glaucoma.

En-face optical coherence tomography - a novel application of non-invasive imaging to art conservation.

Optical Coherence Tomography (OCT) is an optical interferometric technique developed mainly for in vivo imaging of the eye and biological tissues. In this paper, we demonstrate the potential of OCT for non-invasive examination of museum paintings. Two en-face scanning OCT systems operating at 850 nm and 1300 nm were used to produce B-scan and C-scan images at typical working distances of 2 cm. The 3D images produced by the OCT systems show not only the structure of the varnish layer but also the paint layers and underdrawings (preparatory drawings under the paint layers). The highest ever resolution and dynamic range images of underdrawings are presented and for the first time it is possible to find out non-invasively on which layer the underdrawings were drawn.

3-D optical coherence tomography of the laryngeal mucosa.

Laryngeal carcinoma is one of the commonest primary head and neck malignancy and the need for early identification is very important for successful treatment. Outpatient fibreoptic examination of the larynx is unreliable in differentiating benign, pre-malignant and malignant lesions, and therefore surgeons have to rely on biopsies for a definitive diagnosis. This is an invasive procedure requiring general anaesthesia and may have a detrimental effect on the patient's voice. Conventional imaging modalities (ultrasound, computed tomography and magnetic resonance imaging) have a limited resolution and hence cannot give sufficient information on the extent or nature of laryngeal lesions. The aim of our study is to investigate the feasibility of optical coherence tomography (OCT) in imaging the normal larynx, to lay the foundations for an investigation of its ability to differentiate between benign and malignant disease. Ten tissue specimens from normal larynges were imaged with an 850 nm OCT system that was capable of providing both B-scan (longitudinal or cross-section) images as well as C-scan (en-face or images at constant depth). The en-face OCT mode allowed us to reconstruct 3-D OCT images of the tissue examined. Imaged specimens were processed with standard histopathological techniques and sectioned in the plane of the B-scan OCT images. Haematoxylin-eosin stained specimens were compared with the OCT images thus collected. Preliminary results showed good correlation between OCT images and histology sections in normal tissue.

Topography and volume measurements of the optic nerve usingen-face optical coherence tomography.

A special imaging instrument was developed which can acquire optical coherence tomography (OCT) en-face images from the eye fundus, and simultaneously a confocal image. Using this instrument we illustrate for the first time the application of en-face OCT imaging to produce topography and perform area and volume measurements of the optic nerve. The procedure is compared with the topography, area and volume measurements using a confocal scanning laser ophthalmoscope.

Use of optical coherence tomography for assessment of dental caries: quantitative procedure.

A method for quantitative assessment of dental caries using optical coherence tomography (OCT) was demonstrated. Development of caries lesions in 15 bovine teeth, by demineralization in acidic buffer solution, was quantitatively assessed daily for 3 days, using OCT. An OCT system which can collect A-scans (depth versus reflectivity curve), B-scans (longitudinal images) and C-scans (transverse images at constant depth) was used. While the B- and C-scans qualitatively described the lesion detected, the A-scan which showed the depth (mm) resolved reflectivity (dB) of the tooth tissue was used for the quantitative analysis. After a simple normalization procedure to determine the actual depth the light travelled into the tooth tissue, the area (R) under the A-scan was quantified as a measure of the degree of reflectivity of the tissue. The result showed that R (dB mm) decreased with increasing demineralization time. The percentage reflectivity loss (R%) in demineralized tissue, which related to the amount of mineral loss, was also calculated, and it was observed that R% increased with increasing demineralization time. It was concluded that with the above procedure, OCT could quantitatively monitor the mineral changes in a caries lesion on a longitudinal basis.

Unbalanced versus balanced operation in an optical coherence tomography system.

The choice of a balanced optical coherence tomography (OCT) configuration versus an unbalanced OCT configuration with optimized reference-arm attenuation is discussed. The choice depends on the receiver noise, the fiber-end reflection R, and the power to the object. When OCT is used to investigate biological tissue an equivalent R? can be evaluated as the compound reflected light from tissue. In this case an additional parameter has to be considered: the confocal optical sectioning interval of the OCT system.

Three dimensional OCT images from retina and skin.

We demonstrate the functionality of an en-face optical coherence tomography (OCT) system with images from the retina and skin. En-face images collected at different depths are subsequently used to reconstruct a 3D volume of the tissue. The reconstruction allows software inferred OCT longitudinal images at any transversal position in the stack. The position in depth in the stack before creating longitudinal OCT images is also adjustable, offering a valuable guidance tool for exploring the 3D volume of the tissue. This is illustrated by Quick time movies showing either depth or lateral exploration along one of two possible different directions in the stack of transversal OCT images. Sufficient accuracy of the volume rendered is obtained in 20 seconds when the system operates at 2 frames a second. The system, equipped with the 3D rendering feature acts as a valuable diagnostic tool allowing "peeling off" of transversal and longitudinal biologic material to investigate different internal features.

Noise analysis of a combined optical coherence tomograph and a confocal scanning ophthalmoscope.

A fiberized optical coherence tomographic (OCT) system is modified to produce a confocal image similar to that produced by a scanning laser ophthalmoscope. Two possible configurations are presented, one that can deliver either an OCT or a confocal image and another that is capable of producing both the OCT and the confocal images simultaneously. Using the later configuration, we demonstrate such images from the retina in the living eye. The penalty in terms of performance reduction introduced into the optical coherence tomograph when integrated with a confocal receiver and the signal-to-noise ratio analysis for the different confocal receiver configurations are presented.

En-face coherence imaging using galvanometer scanner modulation.

We introduce a novel optical path-modulation technique for a low-coherence interferometric imaging system based on transverse scanning of the target with a galvanometric scanning-mirror pair. The path modulation arises when the beam that is incident upon one of the scanning mirrors does not fall on its axis of rotation. The method is demonstrated by the production of en-face low-coherence images of different objects such as a fiber-optic tip and a human retina invivo .

Transversal and longitudinal images from the retina of the living eye using low coherence reflectometry.

An optical coherence tomography (OCT) system to produce both longitudinal and transversal images of the in vivo human eye is presented. For the first time, OCT transversal images collected from the living eye at 50-µm depth steps show details unobtainable with the state-of-the-art scanning laser ophthalmoscope. Images of up to 3×3 mm are produced from the retina in less than a second. For images larger than 1.6×1.6 mm, a path modulation is introduced by the galvanometric scanning mirror and is used as an effective phase modulation method.© 1998 Society of Photo-Optical Instrumentation Engineers.

Simultaneous en-face imaging of two layers in the human retina by low-coherence reflectometry.

We show that with a fiberized multiple Michelson-interferometer-type configuration, transverse images from several layers in the human eye can be simultaneously obtained. We demonstrate the principle by producing simultaneous 100x100 pixel en-face images of a 4 mmx4 mm region on a postmortem retina for two depth positions 250 mum apart.

Practical implementation of a high-speed multichannel correlator with fiber-optic delays.

We discuss architecture, subsystems, and applications of a multichannel digital correlator we constructed by using a combination of optical and electrical delays. Our system is capable of a 1-ns bin width in normal operation, when a derandomizer is used at the input stage and a 0.25-ns bin width when the derandomizer is bypassed. When switchable delay lines are used, the 16 real-time channels can access a 160-ns delay range, providing up to 160 channels at 1-ns bin width or 640 channels at 0.25-ns bin width.

Coherence imaging by use of a Newton rings sampling function.

We show that, with suitable optics in the arm of a Michelson interferometer, orthogonal galvo-scanning mirrors build a sampling function in the form of Newton rings when the two interferometer arms are matched. Using a low-coherence source, one can obtain transversal depth-resolved images. A fast display procedure using a storage oscilloscope was devised based on this method.