Differential interference microscopy (DIC) is a method to obtain the refractive index distribution of a sample as contrast. It is suitable for biological cells, however, DIC can only obtain 2D images from thin samples. Therefore, we introduce a new imaging method, volumetric differential contrast (VDC) imaging using optical coherence tomography (OCT). This method enables getting 3D differential contrast of thick samples. VDC was designed based on the disperse scatterer model (DSM), a new theoretical model of OCT, and obtains differential contrast by complex numerical processing of OCT signal. DSM represents the sample as a spatially distributed refractive index with dispersed random scatterers, and OCT signal was formulated from this model. VDC uses two complex OCT signals, s1 and s2 at two laterally slightly distant positions, and the final image is defined as Im[s1 s2. This signal forms a spatially differential image of the product of the refractive index distribution and the scatterer density. According to the formulation, the size of the differentiation kernel, corresponding to the shear amount of DIC, is proportional to the defocus of the probe beam and the separation between s1 and s2. This method was validated by an in vitro spheroid sample and an in vivo zebrafish sample, measured by spectral domain OCT with a center wavelength of 830 nm. VDC images were obtained from refocused and defocused signals.
We introduce volumetric differential contrast (VDC) imaging using optical coherence tomography (OCT). This method was designed based on a new theoretical model of OCT, the disperse scatterer model (DSM). VDC gives the differential image of “the product of the refractive index distribution and the scatterer density” through complex numerical processing of OCT signals.
The method was validated by in-vitro and in-vivo samples measured by spectral domain OCT. Differential contrast images with arbitrary shear amount and shear direction were obtained at arbitrary depth positions by a single measurement by numerically applying defocus by holographic signal processing after the signal acquisition.
The zebrafish has shown to be an essential preclinical animal model, especially in the field of oncology. A non-invasive, high-resolution, and three-dimensional imaging modality is required to identify disease related changes in this model organism.
The presented polarization-sensitive Jones matrix optical coherence tomography (JM-OCT) prototype was utilized in three different studies, covering in vivo imaging of wildtype zebrafish, an adult postmortem tumor model and a longitudinal xenograft tumor zebrafish investigation.
This work highlights the potential of JM-OCT as a non-invasive, label-free, and three-dimensional imaging tool for preclinical cancer research based on zebrafish models.
The zebrafish is a valuable animal model in pre-clinical cancer research. Optical coherence tomography (OCT) is a non-invasive optical imaging technique, which provides a label-free and three-dimensional method to investigate the tissue structure. Jones-matrix OCT (JM-OCT) is a functional extension of conventional OCT, to gain additional tissue specific contrast by analyzing the polarization states of the back-scattered light. In this work we present the longitudinal investigation of in vivo wildtype and a tumor xenograft zebrafish model using our JM-OCT prototype. The scattering and depth-resolved polarization properties in control versus tumor regions were analyzed and compared to results obtained from histology.
We demonstrate volumetric phase contrast imaging by using optical coherence tomography (OCT). In general, the randomness of the scatterers’ distribution prohibits the volumetric measurement of a meaningful phase in a scattering mode. Our method uses complex numerical manipulation of an en-face complex OCT and gives a transversally differential phase image similar to a differential interference contrast microscope (DIC). Not like the DIC, our method can arbitrarily select the amount and direction of the shear after the OCT acquisition. In addition it provides DIC-like images at arbitrary depths. This method is validated by using a 840-nm spectral domain OCT system. A zebrafish sample is measured over a 1-mm × 1-mm transversal scanning range.
A polarization sensitive Jones matrix optical coherence tomography (JM-OCT) prototype was utilized to investigate in vivo wildtype zebrafish in different development stages. JM-OCT is a functional extension of conventional intensity-based OCT to visualize polarization properties of samples. We demonstrated simultaneous structural and functional imaging of in vivo wildtype zebrafish at the age of eight-days, one and two-months post-fertilization. The scattering and polarization characteristics of the anatomical features, such as the eye, and the muscles, were investigated and quantified non-invasively and in three-dimensions. Furthermore, the setup was able to visualize dynamic processes like the blood flow in a label-free way.
Significance: The scattering and polarization characteristics of various organs of in vivo wildtype zebrafish in three development stages were investigated using a non-destructive and label-free approach. The presented results showed a promising first step for the usability of Jones-matrix optical coherence tomography (JM-OCT) in zebrafish-based research.
Aim: We aim to visualize and quantify the scatter and polarization signatures of various zebrafish organs for larvae, juvenile, and young adult animals in vivo in a non-invasive and label-free way.
Approach: A custom-built polarization-sensitive JM-OCT setup in combination with a motorized translation stage was utilized to investigate live zebrafish. Depth-resolved scattering (intensity and attenuation coefficient) and polarization (birefringence and degree of polarization uniformity) properties were analyzed. OCT angiography (OCT-A) was utilized to investigate the vasculature label-free and non-destructively.
Results: The scatter and polarization signatures of the zebrafish organs such as the eye, gills, and muscles were investigated. The attenuation coefficient and birefringence changes between 1- and 2-month-old animals were evaluated in selected organs. OCT-A revealed the vasculature of in vivo larvae and juvenile zebrafish in a label-free manner.
Conclusions: JM-OCT offers a rapid, label-free, non-invasive, tissue specific, and three-dimensional imaging tool to investigate in vivo processes in zebrafish in various development stages.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.