Abstract

In a scanning laser microscope detecting fluorescent light from the specimen, the depth-discriminating property of confocal scanning has been used to carry out optical slicing of a thick specimen. The recorded digital images constitute a three-dimensional raster covering a volume of the specimen. The specimen has been visualized in stereo and rotation by making look-through projections of the digital data in different directions. The contrast of the pictures has been enhanced by generating the gradient volume. This permits display of the border surfaces between regions instead of the regions themselves.

© 1985 Optical Society of America

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References

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  1. D. A. Agard, J. W. Sedat, Nature 302, 676 (1983).
    [CrossRef] [PubMed]
  2. M. Minsky, “Microscopy apparatus,” U.S. Patent3,013,467 (December191961).
  3. P. Davidovits, M. D. Egger, Appl. Opt. 10, 1615 (1971).
    [CrossRef] [PubMed]
  4. A. F. Slomba, D. E. Wasserman, G. I. Kaufman, J. F. Nester, J. Assoc. Adv. Med. Instrum. 6, 230 (1972).
    [PubMed]
  5. C. J. R. Sheppard, T. Wilson, Opt. Lett. 3, 115 (1978).
    [CrossRef] [PubMed]
  6. G. J. Brakenhoff, P. Blom, P. Barends, J. Microsc. (Oxford) 117, 219 (1979).
    [CrossRef]
  7. T. Wilson, J. Appl. Phys. 22, 119 (1980).
    [CrossRef]
  8. D. K. Hamilton, T. Wilson, C. J. R. Sheppard, Opt. Lett. 6, 625 (1981).
    [CrossRef] [PubMed]
  9. I. J. Cox, C. J. R. Sheppard, T. Wilson, Optik 60, 391 (1982).
  10. I. J. Cox, J. Microsc. 133, 149 (1984).
    [CrossRef] [PubMed]
  11. C. J. R. Sheppard, D. K. Hamilton, Appl. Opt. 22, 886 (1982).
    [CrossRef]
  12. N. Åslund, K. Carlsson, A. Liljeborg, L. Majlöf, in Proceedings of the Third Scandinavian Conference on Image Analysis (Studentlitteratur, Lund, Sweden, 1983), p. 338.
  13. W. W. Stewart, Cell 14, 714 (1978).
    [CrossRef]
  14. P. E. Danielsson, B. Kruse, B. Gudmundsson, in Proceedings of the Workshop on Picture Data Description and Managements (Institute of Electrical and Electronics Engineers, New York, 1980), p. 275.

1984 (1)

I. J. Cox, J. Microsc. 133, 149 (1984).
[CrossRef] [PubMed]

1983 (1)

D. A. Agard, J. W. Sedat, Nature 302, 676 (1983).
[CrossRef] [PubMed]

1982 (2)

I. J. Cox, C. J. R. Sheppard, T. Wilson, Optik 60, 391 (1982).

C. J. R. Sheppard, D. K. Hamilton, Appl. Opt. 22, 886 (1982).
[CrossRef]

1981 (1)

1980 (1)

T. Wilson, J. Appl. Phys. 22, 119 (1980).
[CrossRef]

1979 (1)

G. J. Brakenhoff, P. Blom, P. Barends, J. Microsc. (Oxford) 117, 219 (1979).
[CrossRef]

1978 (2)

1972 (1)

A. F. Slomba, D. E. Wasserman, G. I. Kaufman, J. F. Nester, J. Assoc. Adv. Med. Instrum. 6, 230 (1972).
[PubMed]

1971 (1)

Agard, D. A.

D. A. Agard, J. W. Sedat, Nature 302, 676 (1983).
[CrossRef] [PubMed]

Åslund, N.

N. Åslund, K. Carlsson, A. Liljeborg, L. Majlöf, in Proceedings of the Third Scandinavian Conference on Image Analysis (Studentlitteratur, Lund, Sweden, 1983), p. 338.

Barends, P.

G. J. Brakenhoff, P. Blom, P. Barends, J. Microsc. (Oxford) 117, 219 (1979).
[CrossRef]

Blom, P.

G. J. Brakenhoff, P. Blom, P. Barends, J. Microsc. (Oxford) 117, 219 (1979).
[CrossRef]

Brakenhoff, G. J.

G. J. Brakenhoff, P. Blom, P. Barends, J. Microsc. (Oxford) 117, 219 (1979).
[CrossRef]

Carlsson, K.

N. Åslund, K. Carlsson, A. Liljeborg, L. Majlöf, in Proceedings of the Third Scandinavian Conference on Image Analysis (Studentlitteratur, Lund, Sweden, 1983), p. 338.

Cox, I. J.

I. J. Cox, J. Microsc. 133, 149 (1984).
[CrossRef] [PubMed]

I. J. Cox, C. J. R. Sheppard, T. Wilson, Optik 60, 391 (1982).

Danielsson, P. E.

P. E. Danielsson, B. Kruse, B. Gudmundsson, in Proceedings of the Workshop on Picture Data Description and Managements (Institute of Electrical and Electronics Engineers, New York, 1980), p. 275.

Davidovits, P.

Egger, M. D.

Gudmundsson, B.

P. E. Danielsson, B. Kruse, B. Gudmundsson, in Proceedings of the Workshop on Picture Data Description and Managements (Institute of Electrical and Electronics Engineers, New York, 1980), p. 275.

Hamilton, D. K.

Kaufman, G. I.

A. F. Slomba, D. E. Wasserman, G. I. Kaufman, J. F. Nester, J. Assoc. Adv. Med. Instrum. 6, 230 (1972).
[PubMed]

Kruse, B.

P. E. Danielsson, B. Kruse, B. Gudmundsson, in Proceedings of the Workshop on Picture Data Description and Managements (Institute of Electrical and Electronics Engineers, New York, 1980), p. 275.

Liljeborg, A.

N. Åslund, K. Carlsson, A. Liljeborg, L. Majlöf, in Proceedings of the Third Scandinavian Conference on Image Analysis (Studentlitteratur, Lund, Sweden, 1983), p. 338.

Majlöf, L.

N. Åslund, K. Carlsson, A. Liljeborg, L. Majlöf, in Proceedings of the Third Scandinavian Conference on Image Analysis (Studentlitteratur, Lund, Sweden, 1983), p. 338.

Minsky, M.

M. Minsky, “Microscopy apparatus,” U.S. Patent3,013,467 (December191961).

Nester, J. F.

A. F. Slomba, D. E. Wasserman, G. I. Kaufman, J. F. Nester, J. Assoc. Adv. Med. Instrum. 6, 230 (1972).
[PubMed]

Sedat, J. W.

D. A. Agard, J. W. Sedat, Nature 302, 676 (1983).
[CrossRef] [PubMed]

Sheppard, C. J. R.

Slomba, A. F.

A. F. Slomba, D. E. Wasserman, G. I. Kaufman, J. F. Nester, J. Assoc. Adv. Med. Instrum. 6, 230 (1972).
[PubMed]

Stewart, W. W.

W. W. Stewart, Cell 14, 714 (1978).
[CrossRef]

Wasserman, D. E.

A. F. Slomba, D. E. Wasserman, G. I. Kaufman, J. F. Nester, J. Assoc. Adv. Med. Instrum. 6, 230 (1972).
[PubMed]

Wilson, T.

Appl. Opt. (2)

Cell (1)

W. W. Stewart, Cell 14, 714 (1978).
[CrossRef]

J. Appl. Phys. (1)

T. Wilson, J. Appl. Phys. 22, 119 (1980).
[CrossRef]

J. Assoc. Adv. Med. Instrum. (1)

A. F. Slomba, D. E. Wasserman, G. I. Kaufman, J. F. Nester, J. Assoc. Adv. Med. Instrum. 6, 230 (1972).
[PubMed]

J. Microsc. (1)

I. J. Cox, J. Microsc. 133, 149 (1984).
[CrossRef] [PubMed]

J. Microsc. (Oxford) (1)

G. J. Brakenhoff, P. Blom, P. Barends, J. Microsc. (Oxford) 117, 219 (1979).
[CrossRef]

Nature (1)

D. A. Agard, J. W. Sedat, Nature 302, 676 (1983).
[CrossRef] [PubMed]

Opt. Lett. (2)

Optik (1)

I. J. Cox, C. J. R. Sheppard, T. Wilson, Optik 60, 391 (1982).

Other (3)

N. Åslund, K. Carlsson, A. Liljeborg, L. Majlöf, in Proceedings of the Third Scandinavian Conference on Image Analysis (Studentlitteratur, Lund, Sweden, 1983), p. 338.

M. Minsky, “Microscopy apparatus,” U.S. Patent3,013,467 (December191961).

P. E. Danielsson, B. Kruse, B. Gudmundsson, in Proceedings of the Workshop on Picture Data Description and Managements (Institute of Electrical and Electronics Engineers, New York, 1980), p. 275.

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Figures (3)

Fig. 1
Fig. 1

Ray paths in a confocal microscope scanner. Light from sections of the specimen located out of focus will largely fall outside the aperture and is thus strongly attenuated. This property makes it possible to slice the specimen optically into thin sections. By combining a number of such sections, one may obtain a 3D image of the specimen.

Fig. 2
Fig. 2

Projection method used to produce stereo pairs from the image data obtained by scanning a number of pictures with different focus settings. The projection rays need not be parallel to the bottom plane, as in the figure. It is thus possible to choose any direction for the axis of rotation.

Fig. 3
Fig. 3

Two stereo pairs produced by the method of Fig. 2. The projection angle between the two images of each pair is 8° Note that the dendrites are almost perfectly oriented in a plane. The axon leaves this plane in a perpendicular direction. (a) and (b) were obtained from the original data volume, (c) and (d) from the gradient volume.

Equations (1)

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q = 1 exp ( γ ρ ) .

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