16
Isometric 3D Imaging of Cellular Samples Using
Optical Projection Tomographic Microscopy
Ryan L. Coe, Qin Miao, Kenny F. Chou
Dept. Bioengineering and Human Photonics Lab, University of Washington, Seattle, WA
Michael G. Meyer
VisionGate Inc., 445 N. Fifth St., Phoenix, AZ
Eric J. Seibel
Dept. Mechanical Engineering and Human Photonics Lab, University of Washington, Seattle, WA
16.1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 582
16.2 Theory Underlying Optical Projection Tomography Microscopy (OPTM) . . . . . . . . . . . . . . . . . 583
16.3 Simulation of OPTM Using Individual Microspheres Representing Single Cells . . . . . . . . . . . 588
16.4 Multi-Modal Imaging of Isolated Cells for 3D Cytological Analysis . . . . . . . . . . . . . . . . . . . . . . 598
16.5 Preliminary Multi-Cellular Imaging for Future Biopsy Diagnosis Using OPTM . . . . . . . . . . . 608
16.6 Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6 14
Acknowledgments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 615
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 616
This chapter describes 3D cellular imaging and analysis using the technique of optical projection to-
mography microscopy (OPTM). The theory, simulation, and method of image formation of OPTM
is similar to x-ray computed tomography (CT), but images cells with light in the optical spectrum.
Instead of rotating the scanner and detector around the object being imaged, a cellular specimen
is rotated while the optical axis is fixed. By capturing a series of projection images around the
specimen, a three-dimensional (3D) image can be reconstructed. Absorption-based stains are still
the mainstay of clinical diagnosis, and thus 3D imaging based upon monochrome absorption is
the primary focus. OPTM could play a role in bringing clinical cytopathology and histopathology
into the third dimension. Advanced 3D visualization techniques and multimodal capabilities are
demonstrated, such as 3D fluorescence imaging in epi-illumination that co-registers with nuclear
chromatin morphology stained with common absorptive stains.
Key words: optical projection tomography (OPT), optical projection tomographic microscopy
(OPTM), C ell-CT
T M
, three dimensional (3D) imaging, bright field absorption, epi-illumination flu-
orescence, cytometry, Mie scattering, lung cancer
581

Get Advanced Biophotonics now with O’Reilly online learning.

O’Reilly members experience live online training, plus books, videos, and digital content from 200+ publishers.