Quantitative Microscopy and Tomography of Live Cells
Wide-field Reflection Phase Microscopy
Although the preliminary work of line-field illumination improved the data acquisition rate to 1 kHz, it is limited to monitor membrane fluctuations or intracellular motion only along one dimension. The first full-field phase sensitive OCT was reported using a swept-source configuration, which required 1024 wavelength encoded images to make a volume image. The acquisition rate was not sufficient to observe cell motion . Yamauchi et al.  developed a full-field time-domain reflection phase microscope based on phase shifting interferometry and captured sectional surface profile of living cells at intervals of 1.25 sec, which is still insufficient for studying cell biomechanics.
Previous wide-field reflection phase measurement techniques were limited in performance as they either used frequency domain detection or time-domain detection with temporal modulation. We are developing a single-shot wide-field low coherence reflection phase microscope based on time-domain optical coherence tomography and off-axis interferometry. The surface profile of the sample under investigation will be determined by taking Hilbert transform of the single-shot two-dimensional interferogram. Since single-shot interferograms will be required to retrieve sample phase, we expect to achieve high-speed (e.g., 1 kHz frames/sec) quantitative phase imaging, which will be extremely useful to measure and characterize plasma membrane fluctuations in eukaryotic cells.
Based on the expected advantages of our full-field reflection phase microscope, the LBRC will undertake three interesting mechanobiological research projects: (i) studying non-thermal plasma membrane fluctuations in nucleated cells; (ii) investigating the role of plasma membrane rheological properties in regulating mechanotransduction processes; and (iii) better quantify the role of the mechanical properties of nuclear membrane in laminopathies.
- Sarunic MV, Weinberg S, Izatt JA. Full-field swept-source phase microscopy. Optics Letters. 31, 1462-4 (2006).
- Yamauchi T, Iwai H, Miwa M, Yamashita Y. Low-coherent quantitative phase microscope for nanometer-scale measurement of living cells morphology. Optics Express. 16, 12227-38 (2008). PMID: 18679500.
- Yaqoob Z, Choi W, Oh S, Lue N, Park Y, Fang-Yen C, Dasari RR, Badizadegan K, Feld MS. Improved phase sensitivity in spectral domain phase microscopy using line-field illumination and self phase-referencing. Optics Express. 17, 10681-7 (2009). PMCID: PMC2844447.