Abstract

We show that the focal modulation microscopy (FMM), which combines a spatial phase modulator with confocal microscopy, results in an improvement in spatial resolution. This technique was introduced to increase imaging depth into tissue and rejection of background from a thick scattering object. A theory for image formation in FMM is presented, and the effects of detecting the in-phase modulated fluorescence signal are discussed. Compared with conventional confocal microscopy, the width of the point-spread function for the in-phase fluorescence signal is improved by 16.4%. When applied to saturable fluorescence, the half-width at half-maximum is improved by 33.6%, 50.0%, and 62.9%, at demodulation frequencies 2ω, 4ω, and 8ω, respectively.

© 2009 Optical Society of America

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References

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  1. X. Deng and M. Gu, Appl. Opt. 42, 3321 (2003).
    [CrossRef] [PubMed]
  2. P. J. Dwyer and C. A. DiMarzio, Opt. Lett. 31, 942 (2006).
    [CrossRef] [PubMed]
  3. C. J. R. Sheppard, W. Gong, and K. Si, Opt. Express 16, 17031 (2008).
    [CrossRef] [PubMed]
  4. K. Si, W. Gong, and C. J. R. Sheppard, Appl. Opt. 48, 810 (2009).
    [CrossRef] [PubMed]
  5. L. K. Wong, M. J. Mandella, G. S. Kino, and T. D. Wang, Opt. Lett. 32, 1674 (2007).
    [CrossRef] [PubMed]
  6. N. G. Chen, C. H. Wong, and C. J. R. Sheppard, Opt. Express 16, 18764 (2008).
    [CrossRef]
  7. T. C. Poon, Opt. Lett. 10, 197 (1985).
    [CrossRef] [PubMed]
  8. E. Rittweger, D. Wildanger, and S. W. Hell, EPL 86, 14001 (2009).
    [CrossRef]
  9. M. G. L. Gustafsson, Proc. Natl. Acad. Sci. USA 102, 13081 (2005).
    [CrossRef] [PubMed]
  10. K. Fujita, M. Kobayashi, S. Kawano, M. Yamanaka, and S. Kawata, Phys. Rev. Lett. 99, 228105 (2007).
    [CrossRef]

2009

E. Rittweger, D. Wildanger, and S. W. Hell, EPL 86, 14001 (2009).
[CrossRef]

K. Si, W. Gong, and C. J. R. Sheppard, Appl. Opt. 48, 810 (2009).
[CrossRef] [PubMed]

2008

2007

K. Fujita, M. Kobayashi, S. Kawano, M. Yamanaka, and S. Kawata, Phys. Rev. Lett. 99, 228105 (2007).
[CrossRef]

L. K. Wong, M. J. Mandella, G. S. Kino, and T. D. Wang, Opt. Lett. 32, 1674 (2007).
[CrossRef] [PubMed]

2006

2005

M. G. L. Gustafsson, Proc. Natl. Acad. Sci. USA 102, 13081 (2005).
[CrossRef] [PubMed]

2003

1985

Chen, N. G.

Deng, X.

DiMarzio, C. A.

Dwyer, P. J.

Fujita, K.

K. Fujita, M. Kobayashi, S. Kawano, M. Yamanaka, and S. Kawata, Phys. Rev. Lett. 99, 228105 (2007).
[CrossRef]

Gong, W.

Gu, M.

Gustafsson, M. G. L.

M. G. L. Gustafsson, Proc. Natl. Acad. Sci. USA 102, 13081 (2005).
[CrossRef] [PubMed]

Hell, S. W.

E. Rittweger, D. Wildanger, and S. W. Hell, EPL 86, 14001 (2009).
[CrossRef]

Kawano, S.

K. Fujita, M. Kobayashi, S. Kawano, M. Yamanaka, and S. Kawata, Phys. Rev. Lett. 99, 228105 (2007).
[CrossRef]

Kawata, S.

K. Fujita, M. Kobayashi, S. Kawano, M. Yamanaka, and S. Kawata, Phys. Rev. Lett. 99, 228105 (2007).
[CrossRef]

Kino, G. S.

Kobayashi, M.

K. Fujita, M. Kobayashi, S. Kawano, M. Yamanaka, and S. Kawata, Phys. Rev. Lett. 99, 228105 (2007).
[CrossRef]

Mandella, M. J.

Poon, T. C.

Rittweger, E.

E. Rittweger, D. Wildanger, and S. W. Hell, EPL 86, 14001 (2009).
[CrossRef]

Sheppard, C. J. R.

Si, K.

Wang, T. D.

Wildanger, D.

E. Rittweger, D. Wildanger, and S. W. Hell, EPL 86, 14001 (2009).
[CrossRef]

Wong, C. H.

Wong, L. K.

Yamanaka, M.

K. Fujita, M. Kobayashi, S. Kawano, M. Yamanaka, and S. Kawata, Phys. Rev. Lett. 99, 228105 (2007).
[CrossRef]

Supplementary Material (3)

» Media 1: AVI (1374 KB)     
» Media 2: AVI (772 KB)     
» Media 3: AVI (824 KB)     

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Figures (4)

Fig. 1
Fig. 1

Schematic diagram of the focal modulation microscope. LBE, laser beam expander; SPM, spatial phase modulator; DM, dichroic mirror; LF, long-pass filter; PMT, photomultiplier tube; L 1 , objective lens; L 2 , collection lens (see Media 1, illumination pattern; Media 2, total detected signal without demodulation; and Media 3, detected ac signal).

Fig. 2
Fig. 2

Intensity image of a point object with a point detector for (a) confocal microscope with two identical circular lenses, (b) confocal microscope with divided apertures (D-shaped apertures), (c) modulation signal in FMM, and (d) in-phase signal in FMM. For (a), (b), and (d) this represents the IPSF.

Fig. 3
Fig. 3

Variations of the integrated intensity of IPFMM, compared with the conventional confocal microscope with circular apertures and with D-shaped apertures, for a point detector.

Fig. 4
Fig. 4

PSFs of (a) saturated fluorescence microscopy, and (b) IPFMM combined with saturated excitation of fluorescence for demodulation frequencies ω, 2 ω , 4 ω , and 8 ω , respectively.

Equations (2)

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I ( v x , v y , u , t ) = | h 1 A ( v x , v y , u ) + h 1 B ( v x , v y , u ) e i 2 δ ω t | 2 [ | ( h 2 v x , v y , u ) | 2 2 D ( v x , v y ) ] ,
I ip = ( h 1 A h 1 B + h 1 A h 1 B ) n ( | h 2 | 2 2 D ) .

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