Abstract

We report a scheme for the detector system of confocal microscopes in which the pinhole and a large-area detector are substituted by a CCD camera. The numerical integration of the intensities acquired by the active pixels emulates the signal passing through the pinhole. We demonstrate the imaging capability and the optical sectioning of the system. Subtractive-imaging confocal microscopy can be implemented in a simple manner, providing superresolution and improving optical sectioning.

© 2012 Optical Society of America

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References

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  1. J. B. Pawley, Handbook of Biological Confocal Microscopy, 3rd ed. (Plenum, 2006).
  2. C. J. R. Sheppard and A. Choudhury, Opt. Acta 24, 1051 (1977).
    [CrossRef]
  3. C. J. R. Sheppard and T. Wilson, Opt. Acta 25, 315 (1978).
    [CrossRef]
  4. T. Wilson, Confocal Microscopy (Academic, 1990).
  5. C. J. R. Sheppard and C. J. Cogswell, J. Mod. Opt. 37, 267 (1990).
    [CrossRef]
  6. M. Barth and E. H. K. Stelzer, Optik 96, 53 (1994).
  7. J. Pawley, M. Blouke, and J. Janesick, Proc. SPIE 2655, 125 (1996).
    [CrossRef]
  8. C. B. Müller and J. Enderlein, Phys. Rev. Lett. 104, 198101 (2010).
    [CrossRef]
  9. T. Wilson and R. Carlini, Appl. Opt. 27, 3791 (1988).
    [CrossRef]
  10. R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley, and T. M. Jovin, Micron 34, 293 (2003).
    [CrossRef]
  11. G. Boyer and V. Sarafis, Optik 112, 177 (2000).
    [CrossRef]
  12. M. Martínez-Corral, M. T. Caballero, C. Ibáñez-López, and V. Sarafis, Micron 34, 313 (2003).
    [CrossRef]
  13. M. Gu and C. J. R. Sheppard, J. Opt. Soc. Am. A 9, 151 (1992).
    [CrossRef]
  14. T. Wilson and R. Carlini, Opt. Lett. 12, 227 (1987).
    [CrossRef]

2010 (1)

C. B. Müller and J. Enderlein, Phys. Rev. Lett. 104, 198101 (2010).
[CrossRef]

2003 (2)

R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley, and T. M. Jovin, Micron 34, 293 (2003).
[CrossRef]

M. Martínez-Corral, M. T. Caballero, C. Ibáñez-López, and V. Sarafis, Micron 34, 313 (2003).
[CrossRef]

2000 (1)

G. Boyer and V. Sarafis, Optik 112, 177 (2000).
[CrossRef]

1996 (1)

J. Pawley, M. Blouke, and J. Janesick, Proc. SPIE 2655, 125 (1996).
[CrossRef]

1994 (1)

M. Barth and E. H. K. Stelzer, Optik 96, 53 (1994).

1992 (1)

1990 (1)

C. J. R. Sheppard and C. J. Cogswell, J. Mod. Opt. 37, 267 (1990).
[CrossRef]

1988 (1)

1987 (1)

1978 (1)

C. J. R. Sheppard and T. Wilson, Opt. Acta 25, 315 (1978).
[CrossRef]

1977 (1)

C. J. R. Sheppard and A. Choudhury, Opt. Acta 24, 1051 (1977).
[CrossRef]

Barth, M.

M. Barth and E. H. K. Stelzer, Optik 96, 53 (1994).

Blouke, M.

J. Pawley, M. Blouke, and J. Janesick, Proc. SPIE 2655, 125 (1996).
[CrossRef]

Boyer, G.

G. Boyer and V. Sarafis, Optik 112, 177 (2000).
[CrossRef]

Caballero, M. T.

M. Martínez-Corral, M. T. Caballero, C. Ibáñez-López, and V. Sarafis, Micron 34, 313 (2003).
[CrossRef]

Carlini, R.

Choudhury, A.

C. J. R. Sheppard and A. Choudhury, Opt. Acta 24, 1051 (1977).
[CrossRef]

Cogswell, C. J.

C. J. R. Sheppard and C. J. Cogswell, J. Mod. Opt. 37, 267 (1990).
[CrossRef]

Enderlein, J.

C. B. Müller and J. Enderlein, Phys. Rev. Lett. 104, 198101 (2010).
[CrossRef]

Gu, M.

Hanley, Q. S.

R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley, and T. M. Jovin, Micron 34, 293 (2003).
[CrossRef]

Heintzmann, R.

R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley, and T. M. Jovin, Micron 34, 293 (2003).
[CrossRef]

Ibáñez-López, C.

M. Martínez-Corral, M. T. Caballero, C. Ibáñez-López, and V. Sarafis, Micron 34, 313 (2003).
[CrossRef]

Janesick, J.

J. Pawley, M. Blouke, and J. Janesick, Proc. SPIE 2655, 125 (1996).
[CrossRef]

Jovin, T. M.

R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley, and T. M. Jovin, Micron 34, 293 (2003).
[CrossRef]

Martínez-Corral, M.

M. Martínez-Corral, M. T. Caballero, C. Ibáñez-López, and V. Sarafis, Micron 34, 313 (2003).
[CrossRef]

Müller, C. B.

C. B. Müller and J. Enderlein, Phys. Rev. Lett. 104, 198101 (2010).
[CrossRef]

Munroe, P.

R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley, and T. M. Jovin, Micron 34, 293 (2003).
[CrossRef]

Nailon, J.

R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley, and T. M. Jovin, Micron 34, 293 (2003).
[CrossRef]

Pawley, J.

J. Pawley, M. Blouke, and J. Janesick, Proc. SPIE 2655, 125 (1996).
[CrossRef]

Pawley, J. B.

J. B. Pawley, Handbook of Biological Confocal Microscopy, 3rd ed. (Plenum, 2006).

Sarafis, V.

M. Martínez-Corral, M. T. Caballero, C. Ibáñez-López, and V. Sarafis, Micron 34, 313 (2003).
[CrossRef]

R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley, and T. M. Jovin, Micron 34, 293 (2003).
[CrossRef]

G. Boyer and V. Sarafis, Optik 112, 177 (2000).
[CrossRef]

Sheppard, C. J. R.

M. Gu and C. J. R. Sheppard, J. Opt. Soc. Am. A 9, 151 (1992).
[CrossRef]

C. J. R. Sheppard and C. J. Cogswell, J. Mod. Opt. 37, 267 (1990).
[CrossRef]

C. J. R. Sheppard and T. Wilson, Opt. Acta 25, 315 (1978).
[CrossRef]

C. J. R. Sheppard and A. Choudhury, Opt. Acta 24, 1051 (1977).
[CrossRef]

Stelzer, E. H. K.

M. Barth and E. H. K. Stelzer, Optik 96, 53 (1994).

Wilson, T.

T. Wilson and R. Carlini, Appl. Opt. 27, 3791 (1988).
[CrossRef]

T. Wilson and R. Carlini, Opt. Lett. 12, 227 (1987).
[CrossRef]

C. J. R. Sheppard and T. Wilson, Opt. Acta 25, 315 (1978).
[CrossRef]

T. Wilson, Confocal Microscopy (Academic, 1990).

Appl. Opt. (1)

J. Mod. Opt. (1)

C. J. R. Sheppard and C. J. Cogswell, J. Mod. Opt. 37, 267 (1990).
[CrossRef]

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

Micron (2)

M. Martínez-Corral, M. T. Caballero, C. Ibáñez-López, and V. Sarafis, Micron 34, 313 (2003).
[CrossRef]

R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley, and T. M. Jovin, Micron 34, 293 (2003).
[CrossRef]

Opt. Acta (2)

C. J. R. Sheppard and A. Choudhury, Opt. Acta 24, 1051 (1977).
[CrossRef]

C. J. R. Sheppard and T. Wilson, Opt. Acta 25, 315 (1978).
[CrossRef]

Opt. Lett. (1)

Optik (2)

M. Barth and E. H. K. Stelzer, Optik 96, 53 (1994).

G. Boyer and V. Sarafis, Optik 112, 177 (2000).
[CrossRef]

Phys. Rev. Lett. (1)

C. B. Müller and J. Enderlein, Phys. Rev. Lett. 104, 198101 (2010).
[CrossRef]

Proc. SPIE (1)

J. Pawley, M. Blouke, and J. Janesick, Proc. SPIE 2655, 125 (1996).
[CrossRef]

Other (2)

T. Wilson, Confocal Microscopy (Academic, 1990).

J. B. Pawley, Handbook of Biological Confocal Microscopy, 3rd ed. (Plenum, 2006).

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Figures (5)

Fig. 1.
Fig. 1.

Schematic of the proposed CLSM.

Fig. 2.
Fig. 2.

(a) Axial response to a flat mirror for different pinhole sizes (labeled in number of pixels). (b) Profile of a microsphere image of 0.1 μm radius.

Fig. 3.
Fig. 3.

1951 USAF Hi-Resolution target. (a) Confocal image acquired with our system. A titled test target is shown in (b) for the confocal images and in (c) for bright-field images.

Fig. 4.
Fig. 4.

Confocal images of an onion skin cell taken with (a) open pinhole and (b) closed pinhole. In (c) the subtractive image obtained from the open pinhole and closed pinhole images is shown.

Fig. 5.
Fig. 5.

(a) Axial response to a flat mirror and (b) microsphere-image profile in the cases of open pinhole, closed pinhole, and the resulting response with the subtraction of both intensities.

Equations (3)

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

Δpinhole=0.5λNAM,
raxial=1.4λNA2.
Isub=IclosedγIopen.

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