Microscopy

Many opal glasses, when exam ined by means of an object-glass of too low a numerical aperture to show the "opal" material as discrete particles, appear of a yel low or brown colour when illu minated with transmitted light. The depth of the colour seen depends on the density of the opal; it is deeper, also, with a narrow cone of illumination than with a cone of wide angle. With dark ground illumination giving a hollow cone much wider than that corresponding to the numerical aperture of the object-glass, the opal glass will appear of a marked blue colour. Such appearances of colour can be taken as definite evidence of the existence of particles in the object, even though the particles cannot be seen as separate discrete particles with the particular object-glass used.

Similar effects are obtained with fossil diatoms but the appear ances may be complicated by diffraction due to the regular struc ture of the diatom. The colours seen, whether by transmitted light or dark-ground illumination, then depend on the fineness of the structure in relation to the numerical aperture of the lens. If the structure is much too fine to be resolved by the lens used, the diatom shows as a yellow or brown object with well-marked out lines when examined with transmitted light, and as a blue or whitish-blue object with dark-ground illumination. If, however, the structure is almost resolvable by the object-glass used, areas showing blues and greens or pinks may appear on the diatom when a transmitted cone of suitable angle is used, the colours seen with wide-angle dark-ground illumination being usually approximately complementary to those seen with transmitted light. These colours are due to the admission of portions of the first order spectra into the object-glass, and are indications of a regular structure which is almost resolvable by the object-glass. The intensity of the colours depends on the medium in which the dia tom is mounted. Strong colours are shown if the diatom is mounted dry (in air) or in realgar, owing to the large difference of refrac tive index between the material of the diatom and air (lower) or realgar (higher). A weak colour is shown if the diatom is mounted in Canada balsam or styrax, both of which have refractive indices not greatly different from that of the diatom itself.

Small objects having rounded contours with smooth faces may give rise to a pearly appearance under suitable conditions of either transmitted or dark-ground illumination. This is not neces sarily an indication of structure, since the distribution of the pearly light can be shown in some instances to be due to the irregular refractions occurring at the surfaces of the objects. Tous les mois starch grains mounted in water show these lenticular refractions very clearly, though there is, in addition, a slight brownish colour discernible on portions of the grains when exam ined with a narrow cone of transmitted light. This brownish colour

is indicative of structure which, in this object, is the result of the growth of one layer over another. The brown colour is very slight compared with that seen on a diatom such as the Pleuro sigma angulatum, even when this is mounted in Canada balsam. This indicates that the differences of refractive index in the starch grain structure are only very slight.

Growth structures frequently give double refractive effects, and many instances of animal and vegetable tissues and fibres are known in which evidence of structure can be revealed by examination with polarized light. The use of polarized light in the study of natural and artificial fibres used in the textile and other industries is common, and this method of examination is employed for distinguishing between different types of fibres as well as for controlling various industrial processes connected with the treatment and manufacture of fibres. This method of illumi nation has considerable diagnostic value in the study of animal and vegetable tissues.

Recognition of Colour.

The colours of stained specimens can be seen quite clearly by transmitted light, but many naturally coloured objects are either so opaque that they appear black by transmitted light or else, with any method of illumination, there is so much general light mixed with the light coming from the coloured object or detail that its precise hue cannot readily be identified. With ordinary dark-ground illumination, surface colours are liable to be masked by light coming through from the lower parts of the object, and the true colours of the surface are not readily perceived. With the vertical illuminator, the difficul ties arising from this cause are avoided, but the surface colours are much diluted by light regularly reflected from the surface of the object. The ring illuminator gives a hollow cone of illumination of wide angle, similar to that used in ordinary dark-ground illumi nation, the only essential difference being that the ring illuminator reflects the light down on to the surface of the object. As a result of this, illumination by means of the ring illuminator gives a better idea of the true surface colours than that obtained with dark-ground illumination, since the amount of light coming from the surface is bright compared with that coming from lower portions of the object. The ring illuminator also shows up surface colours better than the Vertical Illuminator since, with the former, direct, regular reflections from the surface are largely cut out and the colours seen are diluted with comparatively little of the regularly reflected light. For the study of insect colouring and for much analogous work the Lieberkiihn was at one time exten sively used as the best method of illumination; the ring illuminator gives essentially the same type of illumination as the Lieberkiihn.