Quantitative Microscopy and Tomography of Live Cells

Quantitative 3-D Imaging
The refractive index reveals unique aspects of cellular structure, and is important in studies of cell and tissue light scattering, laser trapping of single cells, flow cytometry, total internal reflection microscopy, and other areas involving the interaction of light with cells and tissues. On the other hand, the refractive index can help the adaptive optics and deconvolution techniques to correct sample-induced aberrations and to achieve more improved resolution. To obtain the refractive index of a live biological sample, we have developed quantitative 3-D imaging techniques.

Based on the quantitative phase microscopy using transmission light, we have developed tomographic phase microscopy (TPM) that can image 3-D refractive index map of a live biological sample. We are trying to enhance the resolution and accuracy of refractive index prediction, increasing the observation time with minimal photo-damage.

Incorporating reflection signal, the axial resolution can be enhanced more than twice. The principle of the reflection tomography has been known for a long time, but its application to a phase object has not been reported in the optical regime. By suppressing the noise from multiple optical surfaces in the beam path, we are trying to implement the reflection tomography.

For high throughput applications, we are developing a synthetic aperture tomography. The proof of concept was successfully demonstrated for a sample in translational motion, and we are trying to apply it to a sample in a microfluidic channel, in which defocus and rotation of the sample are expected. The synthetic aperture tomography can be potentially incorporated into flow cytometry, and can thus enable rapid data acquisition for a large number of cells.

• Tomographic phase microscopy
• Synthetic aperture tomography
• Reflection tomographic phase microscopy

1. Choi, W; Fang-Yen, C; Badizadegan, K; Oh, S; Lue, N; Dasari, RR; Feld, MS, "Tomographic phase microscopy", NATURE METHODS, vol. 4 (9), (2007), p 717.
2. Sung, YJ; Choi, W; Fang-Yen, C; Badizadegan, K; Dasari, RR; Feld, MS, "Optical diffraction tomography for high resolution live cell imaging", OPTICS EXPRESS, vol. 17, (2009), p. 266.
3. D. Nahamoo, S. X. Pan, and A. C. Kak, “Synthetic aperture diffraction tomography and its interpolation-free computer implementation,” IEEE Transactions on Sonics and Ultrasonics, Vol. SU-31, July 1984, pp. 218-229.
4. Niyom Lue, Wonshik Choi, Gabriel Popescu, Kamran Badizadegan, Ramachandra R. Dasari, and Michael S. Feld, "Synthetic aperture tomographic phase microscopy for 3D imaging of live cells in translational motion," Opt. Express 16, 16240-16246 (2008).
5. Slaney, M., “Imaging with Diffraction Tomography,” Ph.D. Thesis,. Purdue University, 1985.