Low-power Phase-contrast Microscopy without a Condenser By D. A by atb57993


									      Low-power Phase-contrast Microscopy without a
                              By D. A. KEMPSON
              (From the Bureau of Animal Population, University of Oxford)

  With those low-power objectives in which a phase-plate can be placed near the
back focal plane, phase-contrast microscopy can be obtained without the use of any
condenser. The illuminating annulus is placed in the conjugate focus of the phase-

    N Y microscope may be adapted for phase-contrast microscopy by the
    \ method described by Kempson et al. (1948) and Baker et al. (1949).
 This method has proved to give excellent results with high powers (2 mm.
 immersion and 4 mm.). It has now been found possible to attain still further
 simplification in its application to low powers (e.g. 16 or 18 mm.).
    It will be remembered that the condenser forms an image of the annular
 light-source at a position just below the object, thus producing a secondary
 light-source, which is then imaged by the objective at a focus behind its rear
 lens, where the phase-plate is placed. Owing to the rear focal plane of high-
 power objectives being generally inside the lens-components, and to the
 limitations of space available inside the mount, it is often necessary to place
 the phase-plate from one to two focal lengths of the objective behind (above)
 the rear focal plane. With a low-power objective, however, such as the Leitz
 No. 3 (18 mm. focal length), it was found possible to place the phase-plate as
 close to the rear focal plane as 2-7 mm., which is about one-seventh of the focal
 length of the objective. Consequently, the secondary light-source can be
 placed some considerable distance away from the object (100 to 200 mm.),
 much further than with high powers, where it must be only a few millimetres
 below the object. It naturally follows that under these conditions, the need of
 even a low-power condenser no longer exists, because the actual light source
 can replace the secondary source formed by a condenser.
    Although theoretically this method is applicable to high powers, there are
 practical disadvantages that weigh against its adoption. The main objections
 are (1) the necessity for a very large light-source, and (2) the fact that parts of
 an ordinary substage will prevent light from the very large annulus required
 from reaching the object.
    Very good phase-contrast was achieved by this method with the objective
 mentioned., The illuminating annulus must be placed from 100 to 200 mm.
'from the object (via the mirror). The size of annulus required for any particular
Quarterly Journal Microscopical Science, Vol. 91, part 1, March 1950.
no        D. A. Kempson—Low-power Phase-contrast Microscopy
Position is easily ascertained by examination of the image formed behind the
rear lens (at the position of the phase-plate), with the aid of either a viewing
telescope or a low-power objective placed in the draw-tube. The tolerance in
the position of the illuminating annulus is due to the fact that precise focus of
its image at the phase-plate is not critical, provided that all the direct light
passes through the phase-plate annulus.
   My thanks are due to Dr. John R. Baker for his constructive criticism and
suggestions towards the production of this paper.

KEMPSON, D . A., THOMAS, O. L., and BAKER, J. R., 1948. Quart. J. micr. Sci., 89, 351.
BAKER, J. R., KEMPSON, D . A., and BRUNET, P. C. J., 1949. Ibid., 90, 323.

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