Quantitative oblique back-illumination microscopy (qOBM) is a label-free imaging technique that enables tomographic phase imaging of thick scattering samples with epi-illumination. Here, we propose the use of two forms of functional imaging with qOBM to study tissue and cell cultures. In doing so, we obtain the spatiotemporal and quantitative functional information associated with the phase values extrapolated from qOBM imaging. We have applied this process to study the efficacy of individual immune T cells to kill glioblastoma spheroid cultures in 3D spheroids. Data show that we can effectively distinguish between cell phenotypes and characterize the dynamic motion of these cells in 3D cultures. This work offers a distinct advantage in tracking 3D cellular dynamics in thick tissue as many function imaging modalities are limited to 2D samples. Further, this technology can be expanded to analyze a wide variety of cellular and subcellular dynamics non-invasively in thick tissue.
Phase imaging and fluorescence microscopy provide valuable complementary information, and individually form the basis for a significant portion of the routing biological and biomedical optical imaging performed today. While multimodal phase and fluorescence microscopy has been explored for thin transparent samples to obtain structural information based on the refractive index distribution (with phase contrast) and molecular content (with fluorescence), combining these complementary technologies to study thick samples has been challenging and remains largely unexplored. This work presents the results of a study that combines quantitative phase imaging (QPI) and refractive index (RI) tomography in thick samples—using quantitative oblique back illumination—and bright field fluorescence deconvolution microscopy. The two technologies use a simple bright field microscope configuration with epi-illumination and through-focus z-stack acquisition, along with a deconvolution algorithm, to achieve 3D imaging. Phase and RI information is acquired nearly simultaneously with the fluorescence information with inherent co-registration of the two modalities. In this work, we will present the theoretical underpinning of this multimodal approach, describe the simple multimodal system, and show imaging results of thick tissues, such as labeled mice brains. This multimodal imaging approach could help biologists and clinicians gain a more comprehensive understanding of the tissue’s morphology and molecular composition, and can be widely applied across a number of biological and biomedical disciplines, including neuroscience, pathology, and oncology.
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.