Abstract

We describe a full-field phase-gradient imaging method: asymmetric illumination-based differential phase contrast (AIDPC). Imaging properties of AIDPC are evaluated using the phase-gradient transfer-function approach and elucidated with experimental images of an optical fiber and a histochemical preparation of a skeletal muscle section. In comparison with full-field differential interference contrast, AIDPC does not require phase shifting for quantitative imaging of phase gradient, provides artifact-free images of birefringent specimens, requires shorter camera exposure, and has larger depth of focus. It is amenable to transfer-function engineering, simultaneous fluorescence imaging, and automated live cell imaging.

© 2009 Optical Society of America

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  1. D. K. Hamilton and C. J. R. Sheppard, J. Microsc. 133, 27 (1984).
    [CrossRef]
  2. D. K. Hamilton, C. J. R.Sheppard, and T. Wilson, J. Microsc. 135, 275 (1984).
    [CrossRef]
  3. W. B. Amos, S. Reichelt, D. M. Cattermole, and J. Laufer, J. Microsc. 210, 166 (2003).
    [CrossRef] [PubMed]
  4. M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, J. Microsc. 214, 7 (2004).
    [CrossRef] [PubMed]
  5. M. Shribak, J. LaFountain, D. Biggs, and S. Inoue, J. Biomed. Opt. 13, 014011-10 (2008).
    [CrossRef] [PubMed]
  6. S. B. Mehta and C. J. R. Sheppard, presented at Focus on Microscopy 2008, April 13-16, 2008, Awaji, Japan. www.focusonmicroscopy.org/2008/PDF/089_pdf.
  7. G. W. Ellis, “Edge enhancement of phase phenomena,” U.S. patent 4,255,014 (March 10, 1981).
  8. B. Kachar, Science 227, 766 (1985).
    [CrossRef] [PubMed]
  9. R. Yi, K. K. Chu, and J. Mertz, Opt. Express 14, 5191 (2006).
    [CrossRef] [PubMed]
  10. T. Noda, S. Kawata, and S. Minami, Appl. Opt. 31, 670 (1992).
    [CrossRef] [PubMed]
  11. C. J. R. Sheppard and T. Wilson, J. Opt. Soc. Am. A 3, 755 (1986).
    [CrossRef]
  12. C. J. Cogswell and C. J. R. Sheppard, J. Microsc. 165, 81 (1992).
    [CrossRef]
  13. S. B. Mehta and C. J. R. Sheppard, Opt. Express 16, 19462 (2008).
    [CrossRef] [PubMed]
  14. T. Wilson and C. J. R. Sheppard, Proc. SPIE 232, 203 (1980).
  15. S. B. Mehta and C. J. R. Sheppard, presented at Focus on Microscopy 2009, April 5-8, 2009, Krakow, Poland, www.focusonmicroscopy.org/2009/PDF/396_Mehta.pdf.

2008 (2)

M. Shribak, J. LaFountain, D. Biggs, and S. Inoue, J. Biomed. Opt. 13, 014011-10 (2008).
[CrossRef] [PubMed]

S. B. Mehta and C. J. R. Sheppard, Opt. Express 16, 19462 (2008).
[CrossRef] [PubMed]

2006 (1)

2004 (1)

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, J. Microsc. 214, 7 (2004).
[CrossRef] [PubMed]

2003 (1)

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

1992 (2)

T. Noda, S. Kawata, and S. Minami, Appl. Opt. 31, 670 (1992).
[CrossRef] [PubMed]

C. J. Cogswell and C. J. R. Sheppard, J. Microsc. 165, 81 (1992).
[CrossRef]

1986 (1)

1985 (1)

B. Kachar, Science 227, 766 (1985).
[CrossRef] [PubMed]

1984 (2)

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

D. K. Hamilton, C. J. R.Sheppard, and T. Wilson, J. Microsc. 135, 275 (1984).
[CrossRef]

1980 (1)

T. Wilson and C. J. R. Sheppard, Proc. SPIE 232, 203 (1980).

Amos, W. B.

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

Arnison, M. R.

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, J. Microsc. 214, 7 (2004).
[CrossRef] [PubMed]

Biggs, D.

M. Shribak, J. LaFountain, D. Biggs, and S. Inoue, J. Biomed. Opt. 13, 014011-10 (2008).
[CrossRef] [PubMed]

Cattermole, D. M.

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

Chu, K. K.

Cogswell, C. J.

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, J. Microsc. 214, 7 (2004).
[CrossRef] [PubMed]

C. J. Cogswell and C. J. R. Sheppard, J. Microsc. 165, 81 (1992).
[CrossRef]

Ellis, G. W.

G. W. Ellis, “Edge enhancement of phase phenomena,” U.S. patent 4,255,014 (March 10, 1981).

Hamilton, D. K.

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

D. K. Hamilton, C. J. R.Sheppard, and T. Wilson, J. Microsc. 135, 275 (1984).
[CrossRef]

Inoue, S.

M. Shribak, J. LaFountain, D. Biggs, and S. Inoue, J. Biomed. Opt. 13, 014011-10 (2008).
[CrossRef] [PubMed]

Kachar, B.

B. Kachar, Science 227, 766 (1985).
[CrossRef] [PubMed]

Kawata, S.

LaFountain, J.

M. Shribak, J. LaFountain, D. Biggs, and S. Inoue, J. Biomed. Opt. 13, 014011-10 (2008).
[CrossRef] [PubMed]

Larkin, K. G.

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, J. Microsc. 214, 7 (2004).
[CrossRef] [PubMed]

Laufer, J.

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

Mehta, S. B.

S. B. Mehta and C. J. R. Sheppard, Opt. Express 16, 19462 (2008).
[CrossRef] [PubMed]

S. B. Mehta and C. J. R. Sheppard, presented at Focus on Microscopy 2008, April 13-16, 2008, Awaji, Japan. www.focusonmicroscopy.org/2008/PDF/089_pdf.

S. B. Mehta and C. J. R. Sheppard, presented at Focus on Microscopy 2009, April 5-8, 2009, Krakow, Poland, www.focusonmicroscopy.org/2009/PDF/396_Mehta.pdf.

Mertz, J.

Minami, S.

Noda, T.

Reichelt, S.

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

Sheppard, C. J. R.

S. B. Mehta and C. J. R. Sheppard, Opt. Express 16, 19462 (2008).
[CrossRef] [PubMed]

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, J. Microsc. 214, 7 (2004).
[CrossRef] [PubMed]

C. J. Cogswell and C. J. R. Sheppard, J. Microsc. 165, 81 (1992).
[CrossRef]

C. J. R. Sheppard and T. Wilson, J. Opt. Soc. Am. A 3, 755 (1986).
[CrossRef]

D. K. Hamilton, C. J. R.Sheppard, and T. Wilson, J. Microsc. 135, 275 (1984).
[CrossRef]

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

T. Wilson and C. J. R. Sheppard, Proc. SPIE 232, 203 (1980).

S. B. Mehta and C. J. R. Sheppard, presented at Focus on Microscopy 2008, April 13-16, 2008, Awaji, Japan. www.focusonmicroscopy.org/2008/PDF/089_pdf.

S. B. Mehta and C. J. R. Sheppard, presented at Focus on Microscopy 2009, April 5-8, 2009, Krakow, Poland, www.focusonmicroscopy.org/2009/PDF/396_Mehta.pdf.

Shribak, M.

M. Shribak, J. LaFountain, D. Biggs, and S. Inoue, J. Biomed. Opt. 13, 014011-10 (2008).
[CrossRef] [PubMed]

Smith, N. I.

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, J. Microsc. 214, 7 (2004).
[CrossRef] [PubMed]

Wilson, T.

C. J. R. Sheppard and T. Wilson, J. Opt. Soc. Am. A 3, 755 (1986).
[CrossRef]

D. K. Hamilton, C. J. R.Sheppard, and T. Wilson, J. Microsc. 135, 275 (1984).
[CrossRef]

T. Wilson and C. J. R. Sheppard, Proc. SPIE 232, 203 (1980).

Yi, R.

Appl. Opt. (1)

J. Biomed. Opt. (1)

M. Shribak, J. LaFountain, D. Biggs, and S. Inoue, J. Biomed. Opt. 13, 014011-10 (2008).
[CrossRef] [PubMed]

J. Microsc. (5)

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

D. K. Hamilton, C. J. R.Sheppard, and T. Wilson, J. Microsc. 135, 275 (1984).
[CrossRef]

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

M. R. Arnison, K. G. Larkin, C. J. R. Sheppard, N. I. Smith, and C. J. Cogswell, J. Microsc. 214, 7 (2004).
[CrossRef] [PubMed]

C. J. Cogswell and C. J. R. Sheppard, J. Microsc. 165, 81 (1992).
[CrossRef]

J. Opt. Soc. Am. A (1)

Opt. Express (2)

Proc. SPIE (1)

T. Wilson and C. J. R. Sheppard, Proc. SPIE 232, 203 (1980).

Science (1)

B. Kachar, Science 227, 766 (1985).
[CrossRef] [PubMed]

Other (3)

S. B. Mehta and C. J. R. Sheppard, presented at Focus on Microscopy 2008, April 13-16, 2008, Awaji, Japan. www.focusonmicroscopy.org/2008/PDF/089_pdf.

G. W. Ellis, “Edge enhancement of phase phenomena,” U.S. patent 4,255,014 (March 10, 1981).

S. B. Mehta and C. J. R. Sheppard, presented at Focus on Microscopy 2009, April 5-8, 2009, Krakow, Poland, www.focusonmicroscopy.org/2009/PDF/396_Mehta.pdf.

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Figures (3)

Fig. 1
Fig. 1

Equivalence of a full-field system using a split source (AIDPC) and a scanning system with a split detector (scanning DPC). The light propagates from left to right in the full-field system and from right to left in the scanning system. Components of the scanning system are labeled in italics. f c , f o , and f l are focal lengths of the condenser/collector, objective, and tube lens, respectively. The phase differentiation is assumed to be along the X direction.

Fig. 2
Fig. 2

Experimental comparison of computed PGTFs in (a) with contrast observed in images in (b) and line profiles in (c), for different imaging modes. In the orthogonal direction, imaging properties of all modes are the same as the bright field.

Fig. 3
Fig. 3

AIDPC segregates absorption and phase-gradient information in a birefringent specimen. Images of H&E-stained skeletal muscle section (Carolina biological #313256) with different imaging modes: (a) AIDPC, (b) PSDIC (according to the four-frame algorithm of [4], (c) DIC image with bias π 5 , (d) average ( I R + I L ) 2 , (e) bright field, and (f) crossed polarizers. Images were acquired using 0.9 NA, 40× objective, matched illumination, and white light from a halogen lamp set at 7 V . Scale bar, 2.5 μ m .

Equations (2)

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I DPC = 2 I R I L I R + I L .
C ( m , n ) = P c ( ξ , η ) 2 P o ( ξ m , η n ) 2 d ξ d η ,

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