Tomasz S. Tkaczyk

Texture analysis of optical coherence tomography images: feasibility for tissue classification

Optical coherence tomography (OCT) acquires cross-sectional images of tissue by measuring back-reflected light. Images from in vivo OCT systems typically have a resolution of 10 to 15 mm, and are thus best suited for visualizing structures in the range of tens to hundreds of microns, such as tissue layers or glands. Many normal and abnormal tissues lack visible structures in this size range, so it may appear that OCT is unsuitable for identification of these tissues. However, examination of structure-poor OCT images reveals that they frequently display a characteristic texture that is due to speckle. We evaluated the application of statistical and spectral texture analysis techniques for differentiating tissue types based on the structural and speckle content in OCT images. Excellent correct classification rates were obtained when images had slight visual differences (mouse skin and fat, correct classification rates of 98.5 and 97.3%, respectively), and reasonable rates were obtained with nearly identical-appearing images (normal versus abnormal mouse lung, correct classification rates of 64.0 and 88.6%, respectively). This study shows that texture analysis of OCT images may be capable of differentiating tissue types without reliance on visible structures.

Snapshot Image Mapping Spectrometer (IMS) with high sampling density for hyperspectral microscopy

A snapshot Image Mapping Spectrometer (IMS) with high sampling density is developed for hyperspectral microscopy, measuring a datacube of dimensions 285 × 285 × 60 (x, y, λ). The spatial resolution is ~0.45 µm with a FOV of 100 × 100 µm2. The measured spectrum is from 450 nm to 650 nm and is sampled by 60 spectral channels with average sampling interval ~3.3 nm. The channel’s spectral resolution is ~8nm. The spectral imaging results demonstrate the potential of the IMS for real-time cellular fluorescence imaging.

Compact Image Slicing Spectrometer (ISS) for hyperspectral fluorescence microscopy

An image slicing spectrometer (ISS) for microscopy applications is presented. Its principle is based on the redirecting of image zones by specially organized thin mirrors within a custom fabricated component termed an image slicer. The demonstrated prototype can simultaneously acquire a 140 nm spectral range within its 2D field of view from a single image. The spectral resolution of the system is 5.6 nm. The FOV and spatial resolution of the ISS depend on the selected microscope objective and for the results presented is 45 x 45 microm(2) and 0.45 microm respectively. This proof-of-concept system can be easily improved in the future for higher (both spectral and spatial) resolution imaging. The system requires no scanning and minimal post data processing. In addition, the reflective nature of the image slicer and use of prisms for spectral dispersion make the system light efficient. Both of the above features are highly valuable for real time fluorescent-spectral imaging in biological and diagnostic applications.

Real-time snapshot hyperspectral imaging endoscope

Hyperspectral imaging has tremendous potential to detect important molecular biomarkers of early cancer based on their unique spectral signatures. Several drawbacks have limited its use for in vivo screening applications: most notably the poor temporal and spatial resolution, high expense, and low optical throughput of existing hyperspectral imagers. We present the development of a new real-time hyperspectral endoscope (called the image mapping spectroscopy endoscope) based on an image mapping technique capable of addressing these challenges. The parallel high throughput nature of this technique enables the device to operate at frame rates of 5.2 frames per second while collecting a (x, y, λ) datacube of 350 × 350 × 48. We have successfully imaged tissue in vivo, resolving a vasculature pattern of the lower lip while simultaneously detecting oxy-hemoglobin.

Realization of refractive microoptics through grayscale lithographic patterning of photosensitive hybrid glass.

Refractive microlenses with more than 50 microm sag are fabricated using grayscale lithography. Mechanical assembly features are made simultaneously alongside the microlenses to facilitate high precision assembly of miniature optical systems. The microlens elements are formed using lithographic patterning of photosensitive hybrid sol-gel glass requiring no etch transfer to the substrate material. Grayscale lithography enables the straightforward patterning of aspheric lenses and arbitrary surfaces within the material depth. Lessons learned in the design of a grayscale photomask are described. Characterization of the fabricated lens elements is reported including lens shape, surface quality, and image quality of a complete assembled imaging system.

Snapshot advantage: a review of the light collection improvement for parallel high-dimensional measurement systems.

Abstract. The snapshot advantage is a large increase in light collection efficiency available to high-dimensional measurement systems that avoid filtering and scanning. After discussing this advantage in the context of imaging spectrometry, where the greatest effort towards developing snapshot systems has been made, we describe the types of measurements where it is applicable. We then generalize it to the larger context of high-dimensional measurements, where the advantage increases geometrically with measurement dimensionality.

High numerical aperture microendoscope objective for a fiber confocal reflectance microscope

A disposable high numerical aperture microendoscope objective has been designed, fabricated, and tested for use with a fiber confocal reflectance microscope. The objective uses high precision LIGA fabricated components to integrate imaging components and hydraulic suction lines into a housing that measures only 3.85 mm in outer diameter and 14.65 mm in length. The hydraulics are used to translate tissue through the focal plane for three dimensional imaging. This device is diffraction limited for lambda = 850 nm, has a numerical aperture of 1.0, a field of view of 250 microm, and a working distance of 450 microm. The objective is intended for in vivo imaging of precancerous cells.

Development of image mappers for hyperspectral biomedical imaging applications.

A new design and fabrication method is presented for creating large-format (>100 mirror facets) image mappers for a snapshot hyperspectral biomedical imaging system called an image mapping spectrometer (IMS). To verify this approach a 250 facet image mapper with 25 multiple-tilt angles is designed for a compact IMS that groups the 25 subpupils in a 5 x 5 matrix residing within a single collecting objectives pupil. The image mapper is fabricated by precision diamond raster fly cutting using surface-shaped tools. The individual mirror facets have minimal edge eating, tilt errors of <1 mrad, and an average roughness of 5.4 nm.

Quantitative sectioning and noise analysis for structured illumination microscopy.

Structured illumination (SI) has long been regarded as a nonquantitative technique for obtaining sectioned microscopic images. Its lack of quantitative results has restricted the use of SI sectioning to qualitative imaging experiments, and has also limited researchers’ ability to compare SI against competing sectioning methods such as confocal microscopy. We show how to modify the standard SI sectioning algorithm to make the technique quantitative, and provide formulas for calculating the noise in the sectioned images. The results indicate that, for an illumination source providing the same spatially-integrated photon flux at the object plane, and for the same effective slice thicknesses, SI sectioning can provide higher SNR images than confocal microscopy for an equivalent setup when the modulation contrast exceeds about 0.09.

Snapshot hyperspectral retinal camera with the Image Mapping Spectrometer (IMS)

We present a snapshot hyperspectral retinal camera with the Image Mapping Spectrometer (IMS) for eye imaging applications. The resulting system is capable of simultaneously acquiring 48 spectral channel images in the range 470 nm–650 nm with frame rate at 5.2 fps. The spatial sampling of each measured spectral scene is 350 × 350 pixels. The advantages of this snapshot device are elimination of the eye motion artifacts and pixel misregistration problems in traditional scanning-based hyperspectral retinal cameras, and real-time imaging of oxygen saturation dynamics with sub-second temporal resolution. The spectral imaging performance is demonstrated in a human retinal imaging experiment in vivo. The absorption spectral signatures of oxy-hemoglobin and macular pigments were successfully acquired by using this device.