Abstract

We apply a wide-field quantitative phase microscopy technique based on parallel two-step phase-shifting on-axis interferometry to visualize live biological cells and microorganism dynamics. The parallel on-axis holographic approach is more efficient with camera spatial bandwidth consumption compared to previous off-axis approaches and thus can capture finer sample spatial details, given a limited spatial bandwidth of a specific digital camera. Additionally, due to the parallel acquisition mechanism, the approach is suitable for visualizing rapid dynamic processes, permitting an interferometric acquisition rate equal to the camera frame rate. The method is demonstrated experimentally through phase microscopy of neurons and unicellular microorganisms.

© 2010 Optical Society of America

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  1. L. H. Deng, X. Trepat, J. P. Butler, E. Millet, K. G. Morgan, D. A. Weitz, and J. J. Fredberg, “Fast and slow dynamics of the cytoskeleton,” Nat. Mater. 5, 636–640 (2006).
    [CrossRef] [PubMed]
  2. M. W. Barnett and P. M. Larkman, “The action potential,” Pract. Neurol. 7, 192–197 (2007).
    [PubMed]
  3. D. Bray, Cell Movements (Garland, 1992).
  4. 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]
  5. 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]
  6. N. T. Shaked, M. T. Rinehart, and A. Wax, “Dual-interference-channel quantitative-phase microscopy of live cell dynamics,” Opt. Lett. 34, 767–769 (2009).
    [CrossRef] [PubMed]
  7. N. T. Shaked, J. D. Finan, F. Guilak, and A. Wax, “Quantitative phase microscopy of articular chondrocyte dynamics by wide-field digital interferometry,” J. Biomed. Opt. 15, 010505 (2010).
    [PubMed]
  8. C. P. Brophy, “Effect of intensity error correlation on the computed phase of phase-shifting interferometry,” J. Opt. Soc. Am. A 7, 537–541 (1990).
    [CrossRef]
  9. Y. Awatsuji, A. Fujii, T. Kubota, and O. Matoba, “Parallel three-step phase-shifting digital holography,” Appl. Opt. 45, 2995–3002 (2006).
    [CrossRef] [PubMed]
  10. A. Stern and B. Javidi, “Space-bandwidth conditions for efficient phase-shifting digital holographic microscopy,” J. Opt. Soc. Am. A 25, 736–741 (2008).
    [CrossRef]
  11. P. Guo and A. J. Devaney, “Digital microscopy using phase-shifting digital holography with two reference waves,” Opt. Lett. 29, 857–859 (2004).
    [CrossRef] [PubMed]
  12. X. F. Meng, L. Z. Cai, X. F. Xu, X. L. Yang, X. X. Shen, G. Y. Dong, and Y. R. Wang, “Two-step phase-shifting interferometry and its application in image encryption,” Opt. Lett. 31, 1414–1416 (2006).
    [CrossRef] [PubMed]
  13. Y. Awatsuji, T. Tahara, A. Kaneko, T. Koyama, K. Nishio, S. Ura, T. Kubota, and O. Matoba, “Parallel two-step phase-shifting digital holography,” Appl. Opt. 47, D183–D189(2008).
    [CrossRef] [PubMed]
  14. J.-P. Liu and T.-C. Poon, “Two-step-only quadrature phase-shifting digital holography,” Opt. Lett. 34, 250–252(2009).
    [CrossRef] [PubMed]
  15. P. Ferraro, S. De Nicola, A. Finizio, G. Coppola, S. Grilli, C. Magro, and G. Pierattini, “Compensation of the inherent wave front curvature in digital holographic coherent microscopy for quantitative phase-contrast imaging,” Appl. Opt. 42, 1938–1946 (2003).
    [CrossRef] [PubMed]
  16. N. T. Shaked, Y. Zhu, M. T. Rinehart, and A. Wax, “Two-step-only phase-shifting interferometry with optimized camera bandwidth for microscopy of live cells,” Opt. Express 17, 15585–15591 (2009).
    [CrossRef] [PubMed]
  17. M. D. Ehlers, “Reinsertion or degradation of AMPA receptors determined by activity-dependent endocytic sorting,” Neuron 28, 511–525 (2000).
    [CrossRef]

2010

N. T. Shaked, J. D. Finan, F. Guilak, and A. Wax, “Quantitative phase microscopy of articular chondrocyte dynamics by wide-field digital interferometry,” J. Biomed. Opt. 15, 010505 (2010).
[PubMed]

2009

2008

2007

M. W. Barnett and P. M. Larkman, “The action potential,” Pract. Neurol. 7, 192–197 (2007).
[PubMed]

2006

2005

2004

2003

2000

M. D. Ehlers, “Reinsertion or degradation of AMPA receptors determined by activity-dependent endocytic sorting,” Neuron 28, 511–525 (2000).
[CrossRef]

1990

Awatsuji, Y.

Badizadegan, K.

Barnett, M. W.

M. W. Barnett and P. M. Larkman, “The action potential,” Pract. Neurol. 7, 192–197 (2007).
[PubMed]

Bray, D.

D. Bray, Cell Movements (Garland, 1992).

Brophy, C. P.

Butler, J. P.

L. H. Deng, X. Trepat, J. P. Butler, E. Millet, K. G. Morgan, D. A. Weitz, and J. J. Fredberg, “Fast and slow dynamics of the cytoskeleton,” Nat. Mater. 5, 636–640 (2006).
[CrossRef] [PubMed]

Cai, L. Z.

Colomb, T.

Coppola, G.

Cuche, E.

Dasari, R. R.

De Nicola, S.

Deflores, L. P.

Deng, L. H.

L. H. Deng, X. Trepat, J. P. Butler, E. Millet, K. G. Morgan, D. A. Weitz, and J. J. Fredberg, “Fast and slow dynamics of the cytoskeleton,” Nat. Mater. 5, 636–640 (2006).
[CrossRef] [PubMed]

Depeursinge, C.

Devaney, A. J.

Dong, G. Y.

Ehlers, M. D.

M. D. Ehlers, “Reinsertion or degradation of AMPA receptors determined by activity-dependent endocytic sorting,” Neuron 28, 511–525 (2000).
[CrossRef]

Emery, Y.

Feld, M. S.

Ferraro, P.

Finan, J. D.

N. T. Shaked, J. D. Finan, F. Guilak, and A. Wax, “Quantitative phase microscopy of articular chondrocyte dynamics by wide-field digital interferometry,” J. Biomed. Opt. 15, 010505 (2010).
[PubMed]

Finizio, A.

Fredberg, J. J.

L. H. Deng, X. Trepat, J. P. Butler, E. Millet, K. G. Morgan, D. A. Weitz, and J. J. Fredberg, “Fast and slow dynamics of the cytoskeleton,” Nat. Mater. 5, 636–640 (2006).
[CrossRef] [PubMed]

Fujii, A.

Grilli, S.

Guilak, F.

N. T. Shaked, J. D. Finan, F. Guilak, and A. Wax, “Quantitative phase microscopy of articular chondrocyte dynamics by wide-field digital interferometry,” J. Biomed. Opt. 15, 010505 (2010).
[PubMed]

Guo, P.

Iwai, H.

Javidi, B.

Kaneko, A.

Koyama, T.

Kubota, T.

Larkman, P. M.

M. W. Barnett and P. M. Larkman, “The action potential,” Pract. Neurol. 7, 192–197 (2007).
[PubMed]

Liu, J.-P.

Magistretti, P. J.

Magro, C.

Marquet, P.

Matoba, O.

Meng, X. F.

Millet, E.

L. H. Deng, X. Trepat, J. P. Butler, E. Millet, K. G. Morgan, D. A. Weitz, and J. J. Fredberg, “Fast and slow dynamics of the cytoskeleton,” Nat. Mater. 5, 636–640 (2006).
[CrossRef] [PubMed]

Morgan, K. G.

L. H. Deng, X. Trepat, J. P. Butler, E. Millet, K. G. Morgan, D. A. Weitz, and J. J. Fredberg, “Fast and slow dynamics of the cytoskeleton,” Nat. Mater. 5, 636–640 (2006).
[CrossRef] [PubMed]

Nishio, K.

Pierattini, G.

Poon, T.-C.

Popescu, G.

Rappaz, B.

Rinehart, M. T.

Shaked, N. T.

Shen, X. X.

Stern, A.

Tahara, T.

Trepat, X.

L. H. Deng, X. Trepat, J. P. Butler, E. Millet, K. G. Morgan, D. A. Weitz, and J. J. Fredberg, “Fast and slow dynamics of the cytoskeleton,” Nat. Mater. 5, 636–640 (2006).
[CrossRef] [PubMed]

Ura, S.

Vaughan, J. C.

Wang, Y. R.

Wax, A.

Weitz, D. A.

L. H. Deng, X. Trepat, J. P. Butler, E. Millet, K. G. Morgan, D. A. Weitz, and J. J. Fredberg, “Fast and slow dynamics of the cytoskeleton,” Nat. Mater. 5, 636–640 (2006).
[CrossRef] [PubMed]

Xu, X. F.

Yang, X. L.

Zhu, Y.

Appl. Opt.

J. Biomed. Opt.

N. T. Shaked, J. D. Finan, F. Guilak, and A. Wax, “Quantitative phase microscopy of articular chondrocyte dynamics by wide-field digital interferometry,” J. Biomed. Opt. 15, 010505 (2010).
[PubMed]

J. Opt. Soc. Am. A

Nat. Mater.

L. H. Deng, X. Trepat, J. P. Butler, E. Millet, K. G. Morgan, D. A. Weitz, and J. J. Fredberg, “Fast and slow dynamics of the cytoskeleton,” Nat. Mater. 5, 636–640 (2006).
[CrossRef] [PubMed]

Neuron

M. D. Ehlers, “Reinsertion or degradation of AMPA receptors determined by activity-dependent endocytic sorting,” Neuron 28, 511–525 (2000).
[CrossRef]

Opt. Express

Opt. Lett.

Pract. Neurol.

M. W. Barnett and P. M. Larkman, “The action potential,” Pract. Neurol. 7, 192–197 (2007).
[PubMed]

Other

D. Bray, Cell Movements (Garland, 1992).

Supplementary Material (2)

» Media 1: AVI (3144 KB)     
» Media 2: AVI (12512 KB)     

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Figures (6)

Fig. 1
Fig. 1

PONI microscope for quantitative phase measurements of live cell and microorganism dynamics: BS 1 and BS 2 , beam splitters; M, mirror; λ / 4 , quarter-wave plate; S, sample; A, rectangular aperture; MO, microscope objective; L 0 , L 1 , and L 2 , lenses; WP, Wollaston prism.

Fig. 2
Fig. 2

Schematic representation of the spatial- frequency domain of (a) the off-axis method (acquisition of a single interferogram) along the axis of the off-axis angle, and (b) the proposed PONI method (simultaneous acquisition of two one-axis interferograms). FT: spatial Fourier transform.

Fig. 3
Fig. 3

Reconstructed amplitude images of groups 4–7 of USAF resolution target obtained by (a) the suggested on-axis method, 0.54 × magnification, no binning, and (b) the traditional off-axis method, 1.08 × magnification, 2 × horizontal binning. The off-axis angle is along the vertical dimension [where resolution loss is seen compared to (a)].

Fig. 4
Fig. 4

Reconstructed amplitude images of groups 7, elements 2–6 of USAF resolution target obtained by (a) the suggested on-axis method, 33 × magnification, 8 × 8 binning, and (b) the traditional off-axis method, 66 × magnification, 8 × 8 binning. The off-axis angle is along the vertical dimension [where resolution loss is seen compared to (a)].

Fig. 5
Fig. 5

Rat hippocampal neuron: (a) two phase-shifted on-axis interferograms acquired in a single camera exposure; (b) unwrapped phase profile obtained by the suggested PONI method, 33 × magnification, no binning; (c) unwrapped phase pro file obtained by the traditional off-axis method, 66 × magnification, 2 × vertical binning. The white scale bars indicate 5 μm . Phase colorbar is valid for both (b) and (c). In spite of the fact that (b) is half in size compared to (c), finer details can be detected in (b). Red arrows in (b) indicate two examples for that.

Fig. 6
Fig. 6

Upper part of Euglena gracilis (a unicellular protist): (a) unwrapped phase profile obtained by the suggested PONI method, 33 × magnification, 2 × 2 binning (dynamic behavior: Media 1); (b) unwrapped phase profile obtained by the traditional off-axis method, 66 × magnification, 4 × 2 binning (dynamic behavior: Media 2). The white scale bars indicate 10 μm . Phase colorbar is valid for both (a) and (b).

Equations (5)

Equations on this page are rendered with MathJax. Learn more.

I 1 = I R + I S + 2 I R I S cos ( ϕ OBJ + ϕ C ) ; I 2 = I R + I S + 2 I R I S sin ( ϕ OBJ + ϕ C ) ,
I 0 = I 1 + I 2 + 2 I R ( I 1 + I 2 + 2 I R ) 2 2 ( I 1 2 + I 2 2 + 4 I R 2 ) 2 .
F = [ ( I 1 I S I R ) + j ( I 2 I S I R ) ] / 2 I R ; ϕ ˜ C = arctan ( Im F / Re F ) .
F = [ ( I 1 I 0 ) + j ( I 2 I 0 ) ] / 2 I R ; φ ˜ = arctan ( Im F / Re F ) ,
ϕ OBJ = φ ϕ C .

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