Abstract

We present a method for increasing the lateral resolution and detection efficiency of scanning fluorescence microscopes by adding an interferometer with partial image inversion to the detection pathway. We show that the resulting detection transfer function is essentially the absolute square of the system’s amplitude transfer function enlarged to twice its spatial frequency range. Simulations for a confocal system yield a lateral FWHM resolution of 168 nm (135 nm after image subtraction) as compared to 218 nm for confocal detection without an interferometer. Furthermore we demonstrate how this method is suitable for extended focus imaging. Here simulations for Bessel beam excitation and interferometric detection yield a resolution of 146 nm (116 nm after image subtraction) as compared to 199 nm for integrating detection without an interferometer.

© 2007 Optical Society of America

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  1. A. Egner, S. Verrier, A. Goroshkov, H.-D. Söling and S. Hell, "4Pi-microscopy of the Golgi apparatus in live mammalian cells," J. Struct. Biol. 147, 70-76 (2004).
    [CrossRef]
  2. N. Sandeau and H. Giovannini, "Increasing the lateral resolution of 4Pi fluorescence microscopes," J. Opt. Soc. Am. A 23, 1089-1095 (2006).
    [CrossRef]
  3. N. Sandeau and H. Giovannini, "Arrangement of a 4Pi microscope for reducing the confocal detection volume with two-photon excitation," Opt. Commun. 264, 123-129 (2006).
    [CrossRef]
  4. R. Heintzmann and K. Wicker, UK patent (filed 2 Feb. 2007).
  5. N. Sandeau, H. Rigneault and H. Giovannini, "Increasing the lateral resolution in confocal fluorescence and bio-luminescence microscopes," Abstract book Focus on Microscopy 2007, Valencia, Spain, 248 (2007).
  6. N. Sandeau, H. Rigneault and H. Giovannini, French Patent (filed 8 Jun. 2006).
  7. N. Sandeau and H. Giovannini, "Influence of the pinhole size on the resolution of the 4Pi’ microscope studied by means of the optical transfer function," Nucl. Instrum. Methods Phys. Res. A 571, 404-406 (2007).
    [CrossRef]
  8. B. E. A. Saleh and M. C. Teich, Fundamentals of Photonics, (Wiley-Interscience 1991).
  9. W. Denk, J. H. Strickler and W. W. Webb, "Two-photon laser scanning fluorescence microscopy," Science 248, 73-76 (1990).
    [CrossRef]
  10. R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley and T. M. Jovin, "Resolution enhancement by subtraction of confocal signals taken at different pinhole sizes," Micron. 34, 293-300 (2003).
    [CrossRef]
  11. B. Richards and E. Wolf, "Electromagnetic diffraction in optical systems - II. Structure of the image field in an aplanatic system," Proc. Roy. Soc. 253 A, 358-379 (1959).
  12. J. Durnin, J. J. Miceli, Jr., and J. H. Eberly, "Diffraction-free Beams," Phys. Rev. Lett. 58, 1499-1501 (1987).
    [CrossRef]

2007

N. Sandeau and H. Giovannini, "Influence of the pinhole size on the resolution of the 4Pi’ microscope studied by means of the optical transfer function," Nucl. Instrum. Methods Phys. Res. A 571, 404-406 (2007).
[CrossRef]

2006

N. Sandeau and H. Giovannini, "Arrangement of a 4Pi microscope for reducing the confocal detection volume with two-photon excitation," Opt. Commun. 264, 123-129 (2006).
[CrossRef]

N. Sandeau and H. Giovannini, "Increasing the lateral resolution of 4Pi fluorescence microscopes," J. Opt. Soc. Am. A 23, 1089-1095 (2006).
[CrossRef]

2004

A. Egner, S. Verrier, A. Goroshkov, H.-D. Söling and S. Hell, "4Pi-microscopy of the Golgi apparatus in live mammalian cells," J. Struct. Biol. 147, 70-76 (2004).
[CrossRef]

2003

R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley and T. M. Jovin, "Resolution enhancement by subtraction of confocal signals taken at different pinhole sizes," Micron. 34, 293-300 (2003).
[CrossRef]

1990

W. Denk, J. H. Strickler and W. W. Webb, "Two-photon laser scanning fluorescence microscopy," Science 248, 73-76 (1990).
[CrossRef]

1987

J. Durnin, J. J. Miceli, Jr., and J. H. Eberly, "Diffraction-free Beams," Phys. Rev. Lett. 58, 1499-1501 (1987).
[CrossRef]

J. Opt. Soc. Am. A

J. Struct. Biol.

A. Egner, S. Verrier, A. Goroshkov, H.-D. Söling and S. Hell, "4Pi-microscopy of the Golgi apparatus in live mammalian cells," J. Struct. Biol. 147, 70-76 (2004).
[CrossRef]

Micron.

R. Heintzmann, V. Sarafis, P. Munroe, J. Nailon, Q. S. Hanley and T. M. Jovin, "Resolution enhancement by subtraction of confocal signals taken at different pinhole sizes," Micron. 34, 293-300 (2003).
[CrossRef]

Nucl. Instrum. Methods Phys. Res. A

N. Sandeau and H. Giovannini, "Influence of the pinhole size on the resolution of the 4Pi’ microscope studied by means of the optical transfer function," Nucl. Instrum. Methods Phys. Res. A 571, 404-406 (2007).
[CrossRef]

Opt. Commun.

N. Sandeau and H. Giovannini, "Arrangement of a 4Pi microscope for reducing the confocal detection volume with two-photon excitation," Opt. Commun. 264, 123-129 (2006).
[CrossRef]

Phys. Rev. Lett.

J. Durnin, J. J. Miceli, Jr., and J. H. Eberly, "Diffraction-free Beams," Phys. Rev. Lett. 58, 1499-1501 (1987).
[CrossRef]

Science

W. Denk, J. H. Strickler and W. W. Webb, "Two-photon laser scanning fluorescence microscopy," Science 248, 73-76 (1990).
[CrossRef]

Other

R. Heintzmann and K. Wicker, UK patent (filed 2 Feb. 2007).

N. Sandeau, H. Rigneault and H. Giovannini, "Increasing the lateral resolution in confocal fluorescence and bio-luminescence microscopes," Abstract book Focus on Microscopy 2007, Valencia, Spain, 248 (2007).

N. Sandeau, H. Rigneault and H. Giovannini, French Patent (filed 8 Jun. 2006).

B. E. A. Saleh and M. C. Teich, Fundamentals of Photonics, (Wiley-Interscience 1991).

B. Richards and E. Wolf, "Electromagnetic diffraction in optical systems - II. Structure of the image field in an aplanatic system," Proc. Roy. Soc. 253 A, 358-379 (1959).

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