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