Abstract

We report on cross-sectional imaging of dynamic biological specimens using a spectral domain phase microscopy (SDPM) system capable of operating at a line rate of 19 kHz. This system combines the time-sensitive capabilities of SDPM with the multi-point acquisition features of related phase-sensitive techniques. The presented phase portraits and B-scan phase images of spontaneously beating embryonic cardiomyocytes and cytoplasmic flow in A. proteus offer insight into the nature and timing of the observed cellular phenomena, demonstrating the utility of this technique for dynamic cell studies.

© 2007 Optical Society of America

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    [CrossRef] [PubMed]
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    [CrossRef]
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    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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    [CrossRef]
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    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]

2007 (3)

2006 (4)

2005 (6)

2004 (1)

2003 (3)

1998 (1)

1991 (1)

D. Huang, "Optical Coherence Tomography," Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Appl. Opt. (1)

J. Biomed. Opt. (2)

G. Popescu, T. Ikeda, C. A. Best, K. Badizadegan, R. R. Dasari, and M. S. Feld, "Erythrocyte structure and dynamics quantified by Hilbert phase microscopy," J. Biomed. Opt. 10, 060503 (2005).
[CrossRef]

M. A. Choma, A. K. Ellerbee, S. Yazdanfar, and J. A. Izatt, "Doppler flow imaging of cytoplasmic streaming using spectral domain phase microscopy," J. Biomed. Opt. 11, 024014 (2006).
[CrossRef] [PubMed]

Opt. Express (4)

Opt. Lett. (11)

G. Popescu, L. P. Deflores, J. C. Vaughan, K. Badizadegan, H. Iwai, R. R. Dasari, and M. S. Feld, "Fourier phase microscopy for investigation of biological structures and dynamics," Opt. Lett. 29, 2503-2505 (2004).
[CrossRef] [PubMed]

P. Marquet, B. Rappaz, P. J. Magistretti, E. Cuche, Y. Emery, T. Colomb, and C. Depeursinge, "Digital holographic microscopy: a noninvasive contrast imaging technique allowing quantitative visualization of living cells with subwavelength axial accuracy," Opt. Lett. 30, 468-470 (2005).
[CrossRef] [PubMed]

Y. Park, G. Popescu, K. Badizadegan, R. R. Dasari, and M. S. Feld, "Fresnel particle tracing in three dimensions using diffraction phase microscopy," Opt. Lett. 32, 811 (2007).
[CrossRef] [PubMed]

T. Ikeda, G. Popescu, R. R. Dasari, and M. S. Feld, "Hilbert phase microscopy for investigating fast dynamics in transparent systems," Opt. Lett. 30, 1165-1167 (2005).
[CrossRef] [PubMed]

J. de Boer, B. Cense, B. H. Park, M. C. Pierce, G. J. Tearney, and B. E. Bouma, "Improved signal-to-noise ratio in spectral-domain compared with time-domain optical coherence tomography," Opt. Lett. 28, 2067-2069 (2003).
[CrossRef] [PubMed]

M. A. Choma, A. K. Ellerbee, C. Yang, T. L. Creazzo, and J. A. Izatt, "Spectral-domain phase microscopy," Opt. Lett. 30, 1162-1164 (2005).
[CrossRef] [PubMed]

A. Barty, K. A. Nugent, D. Paganin, and A. Roberts, "Quantitative optical phase microscopy," Opt. Lett. 23, 817-819 (1998).
[CrossRef]

X. Li, T. Yamauchi, H. Iwai, Y. Yamashita, H. Zhang, and T. Hiruma, "Full-field quantitative phase imaging by white-light interferometry with active phase stabilization and its application to biological samples," Opt. Lett. 31, 1830-1832 (2006).
[CrossRef] [PubMed]

M. V. Sarunic, S. Weinberg, and J. A. Izatt, "Full-field swept-source phase microscopy," Opt. Lett. 31, 1462-1464 (2006).
[CrossRef] [PubMed]

C. Joo, T. Akkin, B. Cense, B. H. Park, and J. F. de Boer, "Spectral-domain optical coherence phase microscopy for quantitative phase-contrast imaging," Opt. Lett. 30, 2131-2133 (2005).
[CrossRef] [PubMed]

C. Joo, K. H. Kim, and J. F. de Boer, "Spectral-domain optical coherence phase and multi-photon microscopy," Opt. Lett. 32, 623-625 (2007).
[CrossRef] [PubMed]

Science (1)

D. Huang, "Optical Coherence Tomography," Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Other (2)

E. J. McDowell, A. K. Ellerbee, M. A. Choma, B. E. Applegate, and J. A. Izatt are preparing a manuscript to be called "Spectral domain phase microscopy for local measurements of cytoskeletal rheology in single cells."

V. Hamburger and H. L. Hamilton, "A series of normal stages in the development of the chick embryo," developmental dynamics 195, 231-272 (1992).
[CrossRef] [PubMed]

Supplementary Material (1)

» Media 1: AVI (1987 KB)     

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Figures (4)

Fig. 1.
Fig. 1.

MD-SDPM schematic. The system employs a broadband source (Ti:Sapphire laser, λ0 = 780 nm, Δλ ~ 70–90 nm) and customized spectrometer (19 kHz line rate) that have been fitted to the documentation port of an inverted microscope. Lateral imaging is performed by raster scanning. FC - fiber coupler; L - lens; DP - documentation port; OBJ - objective; TL - tube lens; CS - coverslip; CCD - charge coupled device; θx,y - galvonometer mirror scan pair.

Fig. 2.
Fig. 2.

(a). Amplitude B-scan captured during a contraction event (t = 625 ms) in the isolated cell shown in b. (b) Scan position indicated by red line (10.95 μm). (c) Corresponding phase B-reveals two domains within the cell moving in opposite directions. Color bar denotes relative phase (in radians). Intensity-based mask applied to phase data suppresses noise and emphasizes cell motion. (1.06 MB) movie acquired over the first two beats. (d) Phase traces of two lateral positions (1.97 μm and 6.02 μm, red and blue, respectively) at the same axial depth (44.9 μm) show different motion between the right and left cell domains. [Media 1]

Fig. 3.
Fig. 3.

(a). B-scan, or lateral plane image, of the cell cluster shown in b; (b) Bright field photomicrograph of a ventricular cardiomyocyte cluster from a Stage 14 chick embryo. Red line denotes the position of the scan, which is 52.75 μm in length; (c) Phase portraits corresponding to points from the multi-cell cluster identified in a are marked in white in b. The sampling rate was possibly too low (12.5Hz) to capture any resting period between successive beats, leading to interpretive challenges. Point 1 can be said to be beating out of phase with points 2 or 3, or to be moving in the opposite direction.

Fig. 4.
Fig. 4.

MD-SDPM data of A. Proteus cytoplasmic flow, flow directed into the page. (a) B-scan image of mean amplitude overlaid with the mean Doppler frequency shift over 9 s at each pixel for |f Dopp| > 0.15 Hz. (b) Lateral Doppler frequency and (c) lateral unwrapped phase distribution over time taken from the position corresponding to the dashed horizontal line in a (depth = 48.6 μm). The flow manifests itself as a trend of steadily increasing phase confined within the cell, and is visible in c. (d) Axial M-mode and (e) Doppler frequency distribution over time taken from the position corresponding to the dashed vertical line in a (depth = 74.5 μm).

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