Houssine Makhlouf

Multispectral confocal microendoscope for in vivo and in situ imaging

We describe the design and operation of a multispectral confocal microendoscope. This fiber-based fluorescence imaging system consists of a slit-scan confocal microscope coupled to an imaging catheter that is designed to be minimally invasive and allow for cellular level imaging in vivo. The system can operate in two imaging modes. The grayscale mode of operation provides high resolution real-time in vivo images showing the intensity of fluorescent signal from the specimen. The multispectral mode of operation uses a prism as a dispersive element to collect a full multispectral image of the fluorescence emission. The instrument can switch back and forth nearly instantaneously between the two imaging modes (less than half a second). In the current configuration, the multispectral confocal microendoscope achieves 3-microm lateral resolution and 30-microm axial resolution. The system records light from 500 to 750 nm, and the minimum resolvable wavelength difference varies from 2.9 to 8.3 nm over this spectral range. Grayscale and multispectral imaging results from ex-vivo human tissues and small animal tissues are presented.

Dual modality fluorescence confocal and spectral-domain optical coherence tomography microendoscope

Optical biopsy facilitates in vivo disease diagnoses by providing a real-time in situ view of tissue in a clinical setting. Fluorescence confocal microendoscopy and optical coherence tomography (OCT) are two methods that have demonstrated significant potential in this context. These techniques provide complementary viewpoints. The high resolution and contrast associated with confocal systems allow en face visualization of sub-cellular details and cellular organization within a thin layer of biological tissue. OCT provides cross-sectional images showing the tissue micro-architecture to a depth beyond the reach of confocal systems. We present a novel design for a bench-top imaging system that incorporates both confocal and OCT modalities in the same optical train allowing the potential for rapid switching between the two imaging techniques. Preliminary results using simple phantoms show that it is possible to realize both confocal microendoscopy and OCT through a fiber bundle based imaging system.

Analysis of multimode fiber bundles for endoscopic spectral-domain optical coherence tomography.

A theoretical analysis of the use of a fiber bundle in spectral-domain optical coherence tomography (OCT) systems is presented. The fiber bundle enables a flexible endoscopic design and provides fast, parallelized acquisition of the OCT data. However, the multimode characteristic of the fibers in the fiber bundle affects the depth sensitivity of the imaging system. A description of light interference in a multimode fiber is presented along with numerical simulations and experimental studies to illustrate the theoretical analysis.

Design and Performance of a Multi-Point Scan Confocal Microendoscope

Confocal fluorescence microendoscopy provides high-resolution cellular-level imaging via a minimally invasive procedure, but requires fast scanning to achieve real-time imaging in vivo. Ideal confocal imaging performance is obtained with a point scanning system, but the scan rates required for in vivo biomedical imaging can be difficult to achieve. By scanning a line of illumination in one direction in conjunction with a stationary confocal slit aperture, very high image acquisition speeds can be achieved, but at the cost of a reduction in image quality. Here, the design, implementation, and experimental verification of a custom multi-point aperture modification to a line-scanning multi-spectral confocal microendoscope is presented. This new design improves the axial resolution of a line-scan system while maintaining high imaging rates. In addition, compared to the line-scanning configuration, previously reported simulations predicted that the multi-point aperture geometry greatly reduces the effects of tissue scatter on image quality. Experimental results confirming this prediction are presented.

Integrated fluorescence confocal and spectral-domain optical coherence tomography microendoscope

Optical biopsies are aimed at providing fast and thorough screening of biological tissues in vivo. Disease diagnosis is based on the morphological structures and biochemical features of tissues that can be sampled in situ with high resolution. Some optical screening techniques, such as fluorescence confocal microendoscopy, provide a limited imaging depth due to the shallow penetration of visible light. Despite confocal microendoscopys high resolution and image quality, morphological changes that occur deeper in the tissue cannot be detected. Other imaging techniques, such as optical coherence tomography (OCT), are able to obtain information at greater depth into tissue. A combination of fluorescence confocal and OCT into a single instrument capable of rapidly switching between these modalities, has the potential of providing complementary en face confocal images showing the morphologic features of cells within a surface layer, and cross-sectional OCT images showing tissue microarchitecture below the surface. The concept for this dual system is to utilize the optical train of an existing multi-spectral confocal microendoscope as a spectral-domain OCT system. Progress made on the implementation of this combined dual integrated imaging system is presented. A performance analysis, discussion of the limitations inherent to the use of an imaging fiber bundle, and recent imaging results are presented.

A dual modality fluorescence confocal and optical coherence tomography microendoscope

We demonstrate the implementation of a Fourier domain optical coherence tomography (OCT) imaging system incorporated into the optical train of a fluorescence confocal microendoscope. The slit-scanning confocal system has been presented previously and achieves 3μm lateral resolution and 25μm axial resolution over a field of view of 430μm. Its multi-spectral mode of operation captures images with 6nm average spectral resolution. To incorporate OCT imaging, a common-path interferometer is made with a super luminescent diode and a reference coverslip located at the distal end of the fiber bundle catheter. The infrared diode spectral width allows a theoretical OCT axial resolution of 12.9μm. Light from the reference and sample combine, and a diffraction grating produces a spectral interferogram on the same 2D CCD camera used for confocal microendoscopic imaging. OCT depth information is recovered by a Fourier transform along the spectral dispersion direction. Proper operation of the system scan mirrors allows rapid switching between confocal and OCT imaging modes. The OCT extension takes advantage of the slit geometry, so that a 2D image is acquired without scanning. Combining confocal and OCT imaging modalities provides a more comprehensive view of tissue and the potential to improve disease diagnosis. A preliminary bench-top system design and imaging results are presented.

A multi-point scanner for high frame rate confocal microendoscopy

Slit-scanning geometries for confocal microendoscopy represent a compromise between acquisition rate and optical performance. Such systems provide high frame rates that freeze motion but recent Monte Carlo simulations show that scattered light severely limits the practical imaging depth for in vivo applications. A new multi-point scanning architecture for confocal microendoscopy has been developed. The new scanner is based on a relatively simple modification to the slit-scanning geometry that results in a parallelized point-scanning confocal microendoscope that maintains the high frame rate of a slit-scanning system while providing optical performance close to that of a single point scanning system. The multi-point scanner has been incorporated into an existing multi-spectral slit-scanning confocal microendoscope. The new confocal aperture consists of a slit and a rotating low duty cycle binary transmission grating, which effectively produces a set of continuously moving widely spaced illumination points along the slit. The design maintains the ability to rapidly switch between grayscale and multi-spectral imaging modes. The improved axial resolution of the multi-point scanning confocal microendoscope leads to significantly better confocal sectioning and deeper imaging, which greatly improves the diagnostic potential of the instrument.

Design of a multi-spectral channel for in-vivo confocal microscopy

We present a modified multi-spectral configuration of a slit-scanning confocal microendoscope that provides higher spectral resolution in a fully automated interface. Tissue fluorescence signal is directed through a dispersive element that spreads the spectral information across the CCD camera mapping spectral information perpendicular to the confocal slit. The dispersive element may be chosen to meet the specific requirements defined by the user. Our current system uses a BK7 prism with a 45o wedge angle and a 20mm diameter clear aperture. The prism is shifted from the optical axis allowing automated switching from grayscale (beam on-axis) to multi-spectral (beam off-axis) imaging by tilting a computer controlled mirror. The system records over a spectral range of 450nm to 750nm. The minimum resolvable wavelength difference varies from 2.1nm to 8.3nm over the spectral range. The lateral and axial resolution of the system is approximately 3&mgr;m by 30&mgr;m, respectively, and is the same for both grayscale and multi-spectral imaging modes. Multi-spectral imaging results from ex-vivo tissues are presented.

A Novel Multi-Point Scan Architecture for a High Frame Rate Multi-Spectral Confocal Microendoscope

A new architecture for a confocal microscope is presented. The approach is a hybrid multi-point scanning system that allows real-time grayscale and multi-spectral imaging and is being developed for confocal microendoscopy.

Advances in Microendoscope Design and Application

Significant advances have been made in the design and application of confocal microendoscope systems for in vivo imaging of the human body. This presentation will review progress in the field and highlight important clinical applications.