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A.K. Trull
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In vivo imaging of small animals is of wide interest to the biomedical community studying biological disease and developmental processes. However, optical imaging deep in tissue is severely limited by light scattering, posing restrictions on the imaging depth, image contrast, and spatial resolution. We demonstrate optical coherence projection tomography (OCPT) as a fast three-dimensional optical imaging technique for ballistic, non-scattered light, deep-tissue imaging. OCPT is based on a novel scanning transmission sample arm to rapidly measure ballistic light projections of amplitude and phase through thick biological tissues. We demonstrate the strength of OCPT by imaging an adult zebrafish in a total volume of 1000 mm3 acquired in 24 min. We achieve an unprecedented imaging depth of 4 mm in biological tissue without using optical clearing (up to 27 mean free paths of photon transport). A new way of analyzing optical tomographic imaging depth is demonstrated and applied to OCPT. It shows that the strong light scattering suppression in OCPT is pivotal to reach the SNR limited imaging depth. OCPT allows for a full quantitative assessment of tissue parameters, which is demonstrated by quantifying the attenuation coefficient, refractive index, surface area, and volume of various organs deep inside the zebrafish. Our work opens up the way for longitudinal in vivo small animal studies from the larval to the adult stages.
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In vivo imaging of small animals is of wide interest to the biomedical community studying biological disease and developmental processes. However, optical imaging deep in tissue is severely limited by light scattering, posing restrictions on the imaging depth, image contrast, and spatial resolution. We demonstrate optical coherence projection tomography (OCPT) as a fast three-dimensional optical imaging technique for ballistic, non-scattered light, deep-tissue imaging. OCPT is based on a novel scanning transmission sample arm to rapidly measure ballistic light projections of amplitude and phase through thick biological tissues. We demonstrate the strength of OCPT by imaging an adult zebrafish in a total volume of 1000 mm3 acquired in 24 min. We achieve an unprecedented imaging depth of 4 mm in biological tissue without using optical clearing (up to 27 mean free paths of photon transport). A new way of analyzing optical tomographic imaging depth is demonstrated and applied to OCPT. It shows that the strong light scattering suppression in OCPT is pivotal to reach the SNR limited imaging depth. OCPT allows for a full quantitative assessment of tissue parameters, which is demonstrated by quantifying the attenuation coefficient, refractive index, surface area, and volume of various organs deep inside the zebrafish. Our work opens up the way for longitudinal in vivo small animal studies from the larval to the adult stages.
We present a comparison of image reconstruction techniques for optical projection tomography. We compare conventional filtered back projection, sinogram filtering using the frequency–distance relationship (FDR), image deconvolution, and 2D point-spread-function-based iterative reconstruction. The latter three methods aim to remove the spatial blurring in the reconstructed image originating from the limited depth of field caused by the point spread function of the imaging system. The methods are compared based on simulated data, experimental optical projection tomography data of single fluorescent beads, and high-resolution optical projection tomography imaging of an entire zebrafish larva. We demonstrate that the FDR method performs poorly on data acquired with high numerical aperture optical imaging systems. We show that the deconvolution technique performs best on highly sparse data with low signal-to-noise ratio. The point-spread-function-based reconstruction method is superior for nonsparse objects and data of high signal-to-noise ratio.
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We present a comparison of image reconstruction techniques for optical projection tomography. We compare conventional filtered back projection, sinogram filtering using the frequency–distance relationship (FDR), image deconvolution, and 2D point-spread-function-based iterative reconstruction. The latter three methods aim to remove the spatial blurring in the reconstructed image originating from the limited depth of field caused by the point spread function of the imaging system. The methods are compared based on simulated data, experimental optical projection tomography data of single fluorescent beads, and high-resolution optical projection tomography imaging of an entire zebrafish larva. We demonstrate that the FDR method performs poorly on data acquired with high numerical aperture optical imaging systems. We show that the deconvolution technique performs best on highly sparse data with low signal-to-noise ratio. The point-spread-function-based reconstruction method is superior for nonsparse objects and data of high signal-to-noise ratio.
Disease model systems, such as the zebrafish, play an important role in understanding the onset of diseases like cancer and monitor the efficacy of new drugs. In the past, non-invasive methods for screening, diagnostics and treatment monitoring were intrinsically from the outside. In the past decades, there has been a strong drive to look inside these model systems, which resulted in the development of many small animal tomographic imaging techniques. Due to the absence of ionizing radiation, high-resolution, and cost efficiency, optical tomography is a popular imaging technique to study disease model systems such as zebrafish. The main obstacles in obtaining high-resolution imaging suitable for tissue characterization are the scattering of light in tissue and diffraction of optical waves. Scattering of light in tissue degrades the resolution of optical tomography systems, especially for thick samples. In this thesis, transmission optical coherence tomography (OCT) is used to select ballistic, non-scattered, from non-ballistic, scattered, light. We demonstrate that transmission optical coherence tomography is a versatile tool to measure optical properties of liquids, solids, and particle suspensions. The developed technique is used to perform quantitative optical tomography of the refractive index and attenuation coefficient. A good agreement is observed between our measurements and literature values for group refractive index, group velocity dispersion, and attenuation coefficient. Based on the tomographic reconstruction of transmission OCT measurements, the median attenuation coefficient, group refractive index and volumes of various organs of an adult zebrafish are segmented and quantified in optical coherence projection tomography reconstructions. In optical tomography light is imaged by a lens onto the camera. Due to the focusing of light onto the camera, this light is collected non-uniformly along the propagation direction from the sample. Consequently, the straight-ray assumption as in standard (pre-) clinical X-ray CT reconstruction is violated. Reconstruction of optical tomography images with standard filtered back projection (FBP) causes radial blurring and tangential blurring that becomes stronger with increasing distance to the rotation axis. We present 2D and 3D tomographic reconstruction algorithms that include the point spread function (PSF) of the imaging system. For emission optical projection tomography, these methods show greatly reduced radial and tangential blurring over the entire field of view 113 114 Summary and a significantly improved signal-to-noise ratio compared to FBP. The 3D PSF-based algorithm is evaluated using different initializations. When initialized with the 2D PSF-based reconstruction result, the 3D PSF-based reconstruction gives an improved signal-to-background and image quality in a useful timeframes. Besides including the physical point spread function (PSF) in the 2D tomographic reconstruction, the effect of the PSF also can be reduced by deconvolution of the FBP reconstructed image or filtering the sinogram before FBP reconstruction. We compared the performance of these techniques with each other based on simulations and the signal-to-noise ratio and the sharpness in reconstructed fluorescent beads and zebrafish OPT images. We demonstrate that the sinogram filtering performs poorly on data acquired with high numerical aperture optical imaging systems. We show that the deconvolution technique performs best for highly sparse, low signal-to-noise ratio objects. The PSF-based reconstruction method is superior for non-sparse objects and data of high signal-to-noise ratio. In this thesis, we developed novel algorithms for transmission OCT signal processing and PSF-based tomographic reconstruction. Our algorithms allow for high-resolution quantitative imaging in turbid media. These techniques can be used for quantitative optical imaging of disease model systems. Potentially this may lead to more insight in tissue development and disease onset, progression, and treatment.
...
Disease model systems, such as the zebrafish, play an important role in understanding the onset of diseases like cancer and monitor the efficacy of new drugs. In the past, non-invasive methods for screening, diagnostics and treatment monitoring were intrinsically from the outside. In the past decades, there has been a strong drive to look inside these model systems, which resulted in the development of many small animal tomographic imaging techniques. Due to the absence of ionizing radiation, high-resolution, and cost efficiency, optical tomography is a popular imaging technique to study disease model systems such as zebrafish. The main obstacles in obtaining high-resolution imaging suitable for tissue characterization are the scattering of light in tissue and diffraction of optical waves. Scattering of light in tissue degrades the resolution of optical tomography systems, especially for thick samples. In this thesis, transmission optical coherence tomography (OCT) is used to select ballistic, non-scattered, from non-ballistic, scattered, light. We demonstrate that transmission optical coherence tomography is a versatile tool to measure optical properties of liquids, solids, and particle suspensions. The developed technique is used to perform quantitative optical tomography of the refractive index and attenuation coefficient. A good agreement is observed between our measurements and literature values for group refractive index, group velocity dispersion, and attenuation coefficient. Based on the tomographic reconstruction of transmission OCT measurements, the median attenuation coefficient, group refractive index and volumes of various organs of an adult zebrafish are segmented and quantified in optical coherence projection tomography reconstructions. In optical tomography light is imaged by a lens onto the camera. Due to the focusing of light onto the camera, this light is collected non-uniformly along the propagation direction from the sample. Consequently, the straight-ray assumption as in standard (pre-) clinical X-ray CT reconstruction is violated. Reconstruction of optical tomography images with standard filtered back projection (FBP) causes radial blurring and tangential blurring that becomes stronger with increasing distance to the rotation axis. We present 2D and 3D tomographic reconstruction algorithms that include the point spread function (PSF) of the imaging system. For emission optical projection tomography, these methods show greatly reduced radial and tangential blurring over the entire field of view 113 114 Summary and a significantly improved signal-to-noise ratio compared to FBP. The 3D PSF-based algorithm is evaluated using different initializations. When initialized with the 2D PSF-based reconstruction result, the 3D PSF-based reconstruction gives an improved signal-to-background and image quality in a useful timeframes. Besides including the physical point spread function (PSF) in the 2D tomographic reconstruction, the effect of the PSF also can be reduced by deconvolution of the FBP reconstructed image or filtering the sinogram before FBP reconstruction. We compared the performance of these techniques with each other based on simulations and the signal-to-noise ratio and the sharpness in reconstructed fluorescent beads and zebrafish OPT images. We demonstrate that the sinogram filtering performs poorly on data acquired with high numerical aperture optical imaging systems. We show that the deconvolution technique performs best for highly sparse, low signal-to-noise ratio objects. The PSF-based reconstruction method is superior for non-sparse objects and data of high signal-to-noise ratio. In this thesis, we developed novel algorithms for transmission OCT signal processing and PSF-based tomographic reconstruction. Our algorithms allow for high-resolution quantitative imaging in turbid media. These techniques can be used for quantitative optical imaging of disease model systems. Potentially this may lead to more insight in tissue development and disease onset, progression, and treatment.
Journal article
(2017)
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Anna Trull, Jelle van der Horst, Willem Jan Palenstijn, Lucas van Vliet, Tristan van Leeuwen, Jeroen Kalkman
As a result of the shallow depth of focus of the optical imaging system, the use of standard filtered back projection in optical projection tomography causes space-variant tangential blurring that increases with the distance to the rotation axis. We present a novel optical tomographic image reconstruction technique that incorporates the point spread function of the imaging lens in an iterative reconstruction. The technique is demonstrated using numerical simulations, tested on experimental optical projection tomography data of single fluorescent beads, and applied to high-resolution emission optical projection tomography imaging of an entire zebrafish larva. Compared to filtered back projection our results show greatly reduced radial and tangential blurring over the entire 5.2x5.2 mm2 field of view, and a significantly improved signal to noise ratio.
...
As a result of the shallow depth of focus of the optical imaging system, the use of standard filtered back projection in optical projection tomography causes space-variant tangential blurring that increases with the distance to the rotation axis. We present a novel optical tomographic image reconstruction technique that incorporates the point spread function of the imaging lens in an iterative reconstruction. The technique is demonstrated using numerical simulations, tested on experimental optical projection tomography data of single fluorescent beads, and applied to high-resolution emission optical projection tomography imaging of an entire zebrafish larva. Compared to filtered back projection our results show greatly reduced radial and tangential blurring over the entire 5.2x5.2 mm2 field of view, and a significantly improved signal to noise ratio.
We demonstrate that Fourier-domain transmission OCT is a versatile tool to measure optical material properties of turbid media. We develop an analytical expression for the transmission OCT signal. Based on this analysis we
determine the group refractive index, group velocity dispersion, absorption coecient, and scattering coecient. The optical dispersion is accurately measured for glasses, liquids, and water/glucose mixtures. The optical attenuation is measured in the spatial domain and compared to Mie calculations combined with concentration dependent scattering eects. In the wavevector domain the spectral dependence of the optical attenuation is measured and compared to literature values. The developed technique can be used for optical sensing of
attenuation and dispersion. ...
determine the group refractive index, group velocity dispersion, absorption coecient, and scattering coecient. The optical dispersion is accurately measured for glasses, liquids, and water/glucose mixtures. The optical attenuation is measured in the spatial domain and compared to Mie calculations combined with concentration dependent scattering eects. In the wavevector domain the spectral dependence of the optical attenuation is measured and compared to literature values. The developed technique can be used for optical sensing of
attenuation and dispersion. ...
We demonstrate that Fourier-domain transmission OCT is a versatile tool to measure optical material properties of turbid media. We develop an analytical expression for the transmission OCT signal. Based on this analysis we
determine the group refractive index, group velocity dispersion, absorption coecient, and scattering coecient. The optical dispersion is accurately measured for glasses, liquids, and water/glucose mixtures. The optical attenuation is measured in the spatial domain and compared to Mie calculations combined with concentration dependent scattering eects. In the wavevector domain the spectral dependence of the optical attenuation is measured and compared to literature values. The developed technique can be used for optical sensing of
attenuation and dispersion.
determine the group refractive index, group velocity dispersion, absorption coecient, and scattering coecient. The optical dispersion is accurately measured for glasses, liquids, and water/glucose mixtures. The optical attenuation is measured in the spatial domain and compared to Mie calculations combined with concentration dependent scattering eects. In the wavevector domain the spectral dependence of the optical attenuation is measured and compared to literature values. The developed technique can be used for optical sensing of
attenuation and dispersion.