Abstract

We show the point-spread function of a confocal microscope with an increased detection aperture. The increase in aperture is accomplished by coherent collection of the light from the specimen with two opposing objective lenses, i.e., type-B 4Pi confocal microscopy. We demonstrate experimentally its feasibility for detecting scattered or fluorescently emitted light. The 4Pi confocal point-spread functions are shown for constructive and destructive interference of the collected wave fronts and are compared with the point-spread functions of comparable confocal microscopes.

© 1994 Optical Society of America

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References

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  1. T. Wilson, C. J. R. Sheppard, Theory and Practice of Scanning Optical Microscopy (Academic, London, 1984), Chap. 5, p. 125.
  2. S. Hell, European patent91121368.4 (July2, 1992).
  3. S. Hell, E. H. K Stelzer, J. Opt. Soc. Am. A 9, 2159 (1992).
    [CrossRef]
  4. S. Hell, E. H. K. Stelzer, Opt. Commun. 93, 277 (1992).
    [CrossRef]
  5. M. Born, E. Wolf, Principles of Optics, 6th ed. (Pergamon, Oxford, 1980), Chap. 13, p. 654.
  6. E. Soini, P. Hänninen, S. Hell, Kliin Lab. (Finland) 10, 82 (1993).

1993 (1)

E. Soini, P. Hänninen, S. Hell, Kliin Lab. (Finland) 10, 82 (1993).

1992 (2)

S. Hell, E. H. K Stelzer, J. Opt. Soc. Am. A 9, 2159 (1992).
[CrossRef]

S. Hell, E. H. K. Stelzer, Opt. Commun. 93, 277 (1992).
[CrossRef]

Born, M.

M. Born, E. Wolf, Principles of Optics, 6th ed. (Pergamon, Oxford, 1980), Chap. 13, p. 654.

Hänninen, P.

E. Soini, P. Hänninen, S. Hell, Kliin Lab. (Finland) 10, 82 (1993).

Hell, S.

E. Soini, P. Hänninen, S. Hell, Kliin Lab. (Finland) 10, 82 (1993).

S. Hell, E. H. K Stelzer, J. Opt. Soc. Am. A 9, 2159 (1992).
[CrossRef]

S. Hell, E. H. K. Stelzer, Opt. Commun. 93, 277 (1992).
[CrossRef]

S. Hell, European patent91121368.4 (July2, 1992).

Sheppard, C. J. R.

T. Wilson, C. J. R. Sheppard, Theory and Practice of Scanning Optical Microscopy (Academic, London, 1984), Chap. 5, p. 125.

Soini, E.

E. Soini, P. Hänninen, S. Hell, Kliin Lab. (Finland) 10, 82 (1993).

Stelzer, E. H. K

Stelzer, E. H. K.

S. Hell, E. H. K. Stelzer, Opt. Commun. 93, 277 (1992).
[CrossRef]

Wilson, T.

T. Wilson, C. J. R. Sheppard, Theory and Practice of Scanning Optical Microscopy (Academic, London, 1984), Chap. 5, p. 125.

Wolf, E.

M. Born, E. Wolf, Principles of Optics, 6th ed. (Pergamon, Oxford, 1980), Chap. 13, p. 654.

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

Kliin Lab. (1)

E. Soini, P. Hänninen, S. Hell, Kliin Lab. (Finland) 10, 82 (1993).

Opt. Commun. (1)

S. Hell, E. H. K. Stelzer, Opt. Commun. 93, 277 (1992).
[CrossRef]

Other (3)

M. Born, E. Wolf, Principles of Optics, 6th ed. (Pergamon, Oxford, 1980), Chap. 13, p. 654.

T. Wilson, C. J. R. Sheppard, Theory and Practice of Scanning Optical Microscopy (Academic, London, 1984), Chap. 5, p. 125.

S. Hell, European patent91121368.4 (July2, 1992).

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Figures (3)

Fig. 1
Fig. 1

Setup of the 4Pi(B) confocal microscope: M’s, mirrors; PH’s, pinholes; PMT, photomultiplier tube. Two opposing objective lenses are used to detect coherently the scattered light emanating from the sample.

Fig. 2
Fig. 2

Surface plot of an axial section of the PSF of (a) a confocal and (b), (c) a 4Pi(B) confocal microscope by use of scattered light. Shown is an xz plane containing the optical axis. All PSF’s are normalized. In (b) the path-length difference is a multiple of the wavelength, whereas in (c) it is an odd multiple of half the wavelength. The confocal axial FWHM of 510 nm (a) is reduced to 120 nm for the main maximum in 4Pi(B) confocal microscopy (b). The axial lobes in (b) reach heights of 0.65 and 0.55.

Fig. 3
Fig. 3

The z response for (a) the confocal and (b), (c) the 4Pi(B) confocal fluorescence microscope. All z responses are normalized. In (b) the path-length difference in a multiple of the wavelength, whereas in (c) it is an odd multiple of half the wavelength. We obtained the z responses by scanning a fluorescent layer along the optical axis; these responses represent the laterally integrated PSF. The central maximum in 4Pi(B) confocal microscopy has an axial FWHM of 225 nm (b). The axial lobes in (b) reach heights of 0.73 and 0.76.

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