Abstract

We demonstrate 3D differential phase-contrast (DPC) microscopy, based on computational illumination with a programmable LED array. By capturing intensity images with various illumination angles generated by sequentially patterning an LED array source, we digitally refocus images through various depths via light field processing. The intensity differences from images taken at complementary illumination angles are then used to generate DPC images, which are related to the gradient of phase. The proposed method achieves 3D DPC with simple, inexpensive optics and no moving parts. We experimentally demonstrate our method by imaging a camel hair sample in 3D.

© 2014 Optical Society of America

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    [CrossRef]

2014

2013

2012

T. N. Ford, K. K. Chu, and J. Mertz, Nat. Methods 9, 1195 (2012).
[CrossRef]

L. Waller, G. Situ, and J. Fleischer, Nat. Photonics 6, 474 (2012).
[CrossRef]

2011

2009

S. B. Mehta and C. J. Sheppard, Opt. Lett. 34, 1924 (2009).
[CrossRef]

M. Levoy, Z. Zhang, and I. McDowall, J. Microsc. 235, 144 (2009).
[CrossRef]

2007

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. Dasari, and M. Feld, Nat. Methods 4, 717 (2007).
[CrossRef]

2006

M. Levoy, R. Ng, A. Adams, M. Footer, and M. Horowitz, ACM Trans. Graph. 25, 924 (2006).

2003

W. Amos, S. Reichelt, D. Cattermole, and J. Laufer, J. Microsc. 210, 166 (2003).
[CrossRef]

2001

B. C. Platt and R. Shack, J. Refrac. Surg. 17, S573 (2001).

1984

D. Hamilton and C. Sheppard, J. Microsc. 133, 27 (1984).
[CrossRef]

1967

Adams, A.

M. Levoy, R. Ng, A. Adams, M. Footer, and M. Horowitz, ACM Trans. Graph. 25, 924 (2006).

Amos, W.

W. Amos, S. Reichelt, D. Cattermole, and J. Laufer, J. Microsc. 210, 166 (2003).
[CrossRef]

Andalman, A.

Badizadegan, K.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. Dasari, and M. Feld, Nat. Methods 4, 717 (2007).
[CrossRef]

Broxton, M.

Cattermole, D.

W. Amos, S. Reichelt, D. Cattermole, and J. Laufer, J. Microsc. 210, 166 (2003).
[CrossRef]

Choi, W.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. Dasari, and M. Feld, Nat. Methods 4, 717 (2007).
[CrossRef]

Chu, K. K.

T. N. Ford, K. K. Chu, and J. Mertz, Nat. Methods 9, 1195 (2012).
[CrossRef]

Cohen, N.

Dasari, R.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. Dasari, and M. Feld, Nat. Methods 4, 717 (2007).
[CrossRef]

Deisseroth, K.

Fang-Yen, C.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. Dasari, and M. Feld, Nat. Methods 4, 717 (2007).
[CrossRef]

Feld, M.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. Dasari, and M. Feld, Nat. Methods 4, 717 (2007).
[CrossRef]

Fleischer, J.

L. Waller, G. Situ, and J. Fleischer, Nat. Photonics 6, 474 (2012).
[CrossRef]

Footer, M.

M. Levoy, R. Ng, A. Adams, M. Footer, and M. Horowitz, ACM Trans. Graph. 25, 924 (2006).

Ford, T. N.

J. D. Giese, T. N. Ford, and J. Mertz, Opt. Express 22, 1152 (2014).
[CrossRef]

T. N. Ford, K. K. Chu, and J. Mertz, Nat. Methods 9, 1195 (2012).
[CrossRef]

Frieden, B. R.

Giese, J. D.

Grosenick, L.

Hamilton, D.

D. Hamilton and C. Sheppard, J. Microsc. 133, 27 (1984).
[CrossRef]

Horowitz, M.

M. Levoy, R. Ng, A. Adams, M. Footer, and M. Horowitz, ACM Trans. Graph. 25, 924 (2006).

Horstmeyer, R.

G. Zheng, R. Horstmeyer, and C. Yang, Nat. Photonics 7, 739 (2013).
[CrossRef]

Kolner, C.

Laufer, J.

W. Amos, S. Reichelt, D. Cattermole, and J. Laufer, J. Microsc. 210, 166 (2003).
[CrossRef]

Levoy, M.

M. Broxton, L. Grosenick, S. Yang, N. Cohen, A. Andalman, K. Deisseroth, and M. Levoy, Opt. Express 21, 25418 (2013).
[CrossRef]

M. Levoy, Z. Zhang, and I. McDowall, J. Microsc. 235, 144 (2009).
[CrossRef]

M. Levoy, R. Ng, A. Adams, M. Footer, and M. Horowitz, ACM Trans. Graph. 25, 924 (2006).

Lue, N.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. Dasari, and M. Feld, Nat. Methods 4, 717 (2007).
[CrossRef]

McDowall, I.

M. Levoy, Z. Zhang, and I. McDowall, J. Microsc. 235, 144 (2009).
[CrossRef]

Mehta, S. B.

Mertz, J.

J. D. Giese, T. N. Ford, and J. Mertz, Opt. Express 22, 1152 (2014).
[CrossRef]

T. N. Ford, K. K. Chu, and J. Mertz, Nat. Methods 9, 1195 (2012).
[CrossRef]

Ng, R.

M. Levoy, R. Ng, A. Adams, M. Footer, and M. Horowitz, ACM Trans. Graph. 25, 924 (2006).

Oh, S.

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. Dasari, and M. Feld, Nat. Methods 4, 717 (2007).
[CrossRef]

Platt, B. C.

B. C. Platt and R. Shack, J. Refrac. Surg. 17, S573 (2001).

Reichelt, S.

W. Amos, S. Reichelt, D. Cattermole, and J. Laufer, J. Microsc. 210, 166 (2003).
[CrossRef]

Shack, R.

B. C. Platt and R. Shack, J. Refrac. Surg. 17, S573 (2001).

Sheppard, C.

D. Hamilton and C. Sheppard, J. Microsc. 133, 27 (1984).
[CrossRef]

Sheppard, C. J.

Situ, G.

L. Waller, G. Situ, and J. Fleischer, Nat. Photonics 6, 474 (2012).
[CrossRef]

Waller, L.

L. Waller, G. Situ, and J. Fleischer, Nat. Photonics 6, 474 (2012).
[CrossRef]

Yang, C.

G. Zheng, R. Horstmeyer, and C. Yang, Nat. Photonics 7, 739 (2013).
[CrossRef]

G. Zheng, C. Kolner, and C. Yang, Opt. Lett. 36, 3987 (2011).
[CrossRef]

Yang, S.

Zhang, Z.

M. Levoy, Z. Zhang, and I. McDowall, J. Microsc. 235, 144 (2009).
[CrossRef]

Zheng, G.

G. Zheng, R. Horstmeyer, and C. Yang, Nat. Photonics 7, 739 (2013).
[CrossRef]

G. Zheng, C. Kolner, and C. Yang, Opt. Lett. 36, 3987 (2011).
[CrossRef]

ACM Trans. Graph.

M. Levoy, R. Ng, A. Adams, M. Footer, and M. Horowitz, ACM Trans. Graph. 25, 924 (2006).

J. Microsc.

M. Levoy, Z. Zhang, and I. McDowall, J. Microsc. 235, 144 (2009).
[CrossRef]

D. Hamilton and C. Sheppard, J. Microsc. 133, 27 (1984).
[CrossRef]

W. Amos, S. Reichelt, D. Cattermole, and J. Laufer, J. Microsc. 210, 166 (2003).
[CrossRef]

J. Opt. Soc. Am.

J. Refrac. Surg.

B. C. Platt and R. Shack, J. Refrac. Surg. 17, S573 (2001).

Nat. Methods

T. N. Ford, K. K. Chu, and J. Mertz, Nat. Methods 9, 1195 (2012).
[CrossRef]

W. Choi, C. Fang-Yen, K. Badizadegan, S. Oh, N. Lue, R. Dasari, and M. Feld, Nat. Methods 4, 717 (2007).
[CrossRef]

Nat. Photonics

G. Zheng, R. Horstmeyer, and C. Yang, Nat. Photonics 7, 739 (2013).
[CrossRef]

L. Waller, G. Situ, and J. Fleischer, Nat. Photonics 6, 474 (2012).
[CrossRef]

Opt. Express

Opt. Lett.

Supplementary Material (2)

» Media 1: AVI (2951 KB)     
» Media 2: AVI (6029 KB)     

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Figures (6)

Fig. 1.
Fig. 1.

Experimental setup. A programmable LED array is placed at the back focal plane of the condenser. Each LED generates illumination at a different oblique angle, corresponding to its spatial location in the array.

Fig. 2.
Fig. 2.

(a) A fishscale sample is imaged with all the brightfield LEDs on and then with only (b) the left-half LEDs and (c) the right-half LEDs on. (d) The left–right DPC image is computed from (a) and (b). (e) The top–bottom DPC image is obtained by repeating the same process on images taken with top and bottom halves on.

Fig. 3.
Fig. 3.

3D camel hair sample is illuminated by different LEDs corresponding to different illumination angles. To synthesize intensity at a different focus plane, each image is shifted by an amount proportional to the illumination angle. The shift-and-add process shears the 4D matrix, then integrates across all angles. In order to get DPC images for each synthesized focal plane, we separately add the images corresponding to left and right sides of the source plane. [Online: movie shows images for each brightfield LED (Media 1)]

Fig. 4.
Fig. 4.

Experimental results for 3D DPC. (a), (d), (g), (j), (m) Digitally refocused intensities. (b), (e), (h), (k), (n) Left–right DPC images. (c), (f), (i), (l), (o) Top–bottom DPC images at (a)–(c) z=32μm, (d)–(f) z=12μm, (g)–(i) z=8μm, and (j)–(l) z=42μm. (m)–(o) Magnified images from regions within the dotted orange rectangles. [Online: movies of 3D slices of intensity and DPC images from 100μm to 100 μm with 3 μm increment (Media 2).]

Fig. 5.
Fig. 5.

Images taken with increasing radii of LED illumination corresponding to increased σ from almost zero to one. The features indicated by the red arrows appear in focus with large coherence, but blur out as more LEDs are turned on, indicating better optical sectioning.

Fig. 6.
Fig. 6.

Lateral resolution degrades with increased digital defocus distance, due to unaccounted for broadening of the PSF in our algorithm. We plot here the theoretically achievable lateral resolution as a function of the distance between the synthetic focus plane and the actual focus plane for typical microscope objective properties.

Equations (3)

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

IDPC=(ILIR)/Itot,
ILΔz=left half of bright field LEDsIiΔz,
IDPCΔz=(ILΔzIRΔz)/ItotΔz,

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