Abstract

There are several ways to realize dark-field imaging in confocal microscopy. In a recent paper [J. Microsc. 181 260–268 (1996)] we suggested a simple modification of a commercial confocal microscope to incorporate dark-field imaging. This modification involved an aperture stop covering half of the entrance pupil of the objective lens. Now we investigate the lateral misalignment of the aperture stop for dark-field and stereoscopic confocal microscopes. We show the effect of lateral alignment of the half-stop on the point-spread and transfer functions and also examine the detected signal from a sloping plane reflector. Lateral and axial resolution values are given from theoretical data.

© 1996 Optical Society of America

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References

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  1. P. Török, Z. Laczik, J. N. Skepper, “Simple modification of a commercial scanning laser microscope to incorporate dark-field imaging,” J. Microsc. 181, 260–269 (1996).
    [CrossRef] [PubMed]
  2. P. Török, Z. Laczik, C. J. R. Sheppard, J. N. Skepper, “Dark-field and differential phase contrast imaging modes in confocal microscopy using a half-aperture stop,” presented at the Conference on 3D Imaging Sciences in Microscopy, Oxford, England, April 1996.
  3. C. J. R. Sheppard, D. K. Hamilton, “High resolution stereoscopic imaging,” Appl. Opt. 22, 866–867 (1983).
    [CrossRef]
  4. T. Wilson, C. J. R. Sheppard, Theory and Practice of Scanning Optical Microscopy (Academic, London, 1984).
  5. A. R. Carlini, “Imaging modes of scanning confocal microscopy,” D. Phil. dissertation (University of Oxford, Oxford, UK, 1988).

1996 (1)

P. Török, Z. Laczik, J. N. Skepper, “Simple modification of a commercial scanning laser microscope to incorporate dark-field imaging,” J. Microsc. 181, 260–269 (1996).
[CrossRef] [PubMed]

1983 (1)

C. J. R. Sheppard, D. K. Hamilton, “High resolution stereoscopic imaging,” Appl. Opt. 22, 866–867 (1983).
[CrossRef]

Carlini, A. R.

A. R. Carlini, “Imaging modes of scanning confocal microscopy,” D. Phil. dissertation (University of Oxford, Oxford, UK, 1988).

Hamilton, D. K.

C. J. R. Sheppard, D. K. Hamilton, “High resolution stereoscopic imaging,” Appl. Opt. 22, 866–867 (1983).
[CrossRef]

Laczik, Z.

P. Török, Z. Laczik, J. N. Skepper, “Simple modification of a commercial scanning laser microscope to incorporate dark-field imaging,” J. Microsc. 181, 260–269 (1996).
[CrossRef] [PubMed]

P. Török, Z. Laczik, C. J. R. Sheppard, J. N. Skepper, “Dark-field and differential phase contrast imaging modes in confocal microscopy using a half-aperture stop,” presented at the Conference on 3D Imaging Sciences in Microscopy, Oxford, England, April 1996.

Sheppard, C. J. R.

C. J. R. Sheppard, D. K. Hamilton, “High resolution stereoscopic imaging,” Appl. Opt. 22, 866–867 (1983).
[CrossRef]

P. Török, Z. Laczik, C. J. R. Sheppard, J. N. Skepper, “Dark-field and differential phase contrast imaging modes in confocal microscopy using a half-aperture stop,” presented at the Conference on 3D Imaging Sciences in Microscopy, Oxford, England, April 1996.

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

Skepper, J. N.

P. Török, Z. Laczik, J. N. Skepper, “Simple modification of a commercial scanning laser microscope to incorporate dark-field imaging,” J. Microsc. 181, 260–269 (1996).
[CrossRef] [PubMed]

P. Török, Z. Laczik, C. J. R. Sheppard, J. N. Skepper, “Dark-field and differential phase contrast imaging modes in confocal microscopy using a half-aperture stop,” presented at the Conference on 3D Imaging Sciences in Microscopy, Oxford, England, April 1996.

Török, P.

P. Török, Z. Laczik, J. N. Skepper, “Simple modification of a commercial scanning laser microscope to incorporate dark-field imaging,” J. Microsc. 181, 260–269 (1996).
[CrossRef] [PubMed]

P. Török, Z. Laczik, C. J. R. Sheppard, J. N. Skepper, “Dark-field and differential phase contrast imaging modes in confocal microscopy using a half-aperture stop,” presented at the Conference on 3D Imaging Sciences in Microscopy, Oxford, England, April 1996.

Wilson, T.

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

Appl. Opt. (1)

C. J. R. Sheppard, D. K. Hamilton, “High resolution stereoscopic imaging,” Appl. Opt. 22, 866–867 (1983).
[CrossRef]

J. Microsc. (1)

P. Török, Z. Laczik, J. N. Skepper, “Simple modification of a commercial scanning laser microscope to incorporate dark-field imaging,” J. Microsc. 181, 260–269 (1996).
[CrossRef] [PubMed]

Other (3)

P. Török, Z. Laczik, C. J. R. Sheppard, J. N. Skepper, “Dark-field and differential phase contrast imaging modes in confocal microscopy using a half-aperture stop,” presented at the Conference on 3D Imaging Sciences in Microscopy, Oxford, England, April 1996.

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

A. R. Carlini, “Imaging modes of scanning confocal microscopy,” D. Phil. dissertation (University of Oxford, Oxford, UK, 1988).

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Figures (16)

Fig. 1
Fig. 1

Schematic diagram of the optical system.

Fig. 2
Fig. 2

Absolute value of the confocal PSF’s with parameter ∊. Note the difference between 0° and 90° meridional planes. Contours of 20 equal function values are plotted on a linear scale, and the distributions are normalized to unity. The images correspond to an aperture stop position of ∊ = −0.8.

Fig. 3
Fig. 3

Same as Fig. 2 except for an aperture stop position of ∊ = −0.6.

Fig. 4
Fig. 4

Same as Fig. 2 except for an aperture stop position of ∊ = −0.4.

Fig. 5
Fig. 5

Same as Fig. 2 except for an aperture stop position of ∊ = −0.2.

Fig. 6
Fig. 6

Same as Fig. 2 except for an aperture stop position of ∊ = 0.

Fig. 7
Fig. 7

Same as Fig. 2 except for an aperture stop position of ∊ = 0.2.

Fig. 8
Fig. 8

Same as Fig. 2 except for an aperture stop position of ∊ = 0.4.

Fig. 9
Fig. 9

Angle (β) of the major axis of the FWHM contour ellipse with respect to the optical axis and as a function of aperture stop position ∊.

Fig. 10
Fig. 10

(a) Lateral and (b) axial FWHM values of the overall intensity PSF of aperture stop position ∊. Individual curves correspond to values computed along the 0° and 90° meridional planes. The dot–dash curve corresponds to the size of the minor axis in the 90° plane.

Fig. 11
Fig. 11

Intensity detected from a plane reflector and an on-axis point object as a function of aperture stop position ∊. In-focus calculations were used.

Fig. 12
Fig. 12

Overall transfer function for u = 0. Note the purely real C DF (m, u = 0, ∊).

Fig. 13
Fig. 13

Real and imaginary parts of the overall transfer function for u = 4.

Fig. 14
Fig. 14

Same as Fig. 13 except for u = 8.

Fig. 15
Fig. 15

Detected intensity from a sloping plane reflector. The surface normal angle ϑ of the plane is measured from the optical axis positive direction. The individual images in (a)–(d) correspond to defocus positions of u = 0, 2, 4, and 6.

Fig. 16
Fig. 16

Normalized intensity from a perpendicular plane reflector as a function of defocus. The aperture stop position ∊ is the parameter. When ∊ = 0, the signal vanishes for any u.

Equations (8)

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υ = 2 π λ r sin α u = 8 π λ z 2 sin 2 ( α 2 ) ,
r = ( x 2 2 + y 2 2 ) ( 1 / 2 ) ,
h 1 [ u , υ ; ( 0 ) ] = h 2 [ u , υ ; ( 0 ) ] = π + cos 1 ( ) π cos 1 ( ) d θ | cos θ | 1 d ϱ exp ( 1 2 i u ϱ 2 ) × exp [ i υ ϱ cos ( θ ϕ ) ] ϱ,
h 1 [ u , υ ; ( < 0 ) ] = h 2 [ u , υ ; ( < 0 ) ] = 0 1 exp [ 1 2 i u ϱ 2 ] J 0 ( υ ϱ ) ϱdϱ cos 1 ( ) cos 1 ( ) d θ cos θ 1 d ϱ exp [ 1 2 i u ϱ 2 ] × exp [ i υ ϱ cos ( θ ϕ ) ] ϱ,
h ( u , υ ; ) = h 1 ( u , υ ; ) h 2 ( u , υ ; ) .
c D F ( m , u ; ) = { c ( m , u ) 1 m 1 2 + 2 c ( m , u ) 2 a ( m , u ) 1 2 + 2 > m 0 > m 1 ,
a ( m , u ) = 2 π exp ( i 2 u m 2 ) 0 cos 1 ( 2 m ) 2 m / cos ϑ 1 × exp [ i u ( r m cos ϑ ) 2 ] r × exp ( i u m 2 cos 2 ϑ ) d ϑ d r ,
I ( ϑ , u ; ) = c ( sin ϑ sin α , u ; ) c * ( sin ϑ sin α , u ; ) ,

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