The Microscope. Its History, Construction, and Application 15th ed. / Being a familiar introduction to the use of the instrument, and the study of microscopical science

by Jabez Hogg

Appendix A.

ILLUMINATION ARRANGEMENTS OF THE MICROSCOPE.

A doubt has of late been expressed among practical microscopists as to the value of the illumination arrangements of the lamp and the microscope, so as to secure the more perfect definition of the flagellate organ of the monas and other minute forms of infusorial life. We have been told that better results will be obtained by turning the mirror aside, and so disposing the microscope and lamp in the horizontal position, that the central rays of light from the mirror-edge of the lamp-flame shall pass through the optical axis of the achromatic condenser, the focus of which must be accurately brought upon the field of view by means of the substage centring screws and rack-work, and in such a manner, that by employing a 1-inch objective, a sharply-defined image of the lamp-flame, edge-on, is projected on to the centre of the field in association with the specimen under examination. If the 1-inch objective be now replaced by a 1-12th or 1-16th inch immersion and once again focussed into place, and a slight re-adjustment of the centring made, it will be found that the field is brilliantly illuminated, and the most minute portions of infusorial life are well defined, and with a sharpness otherwise unattainable. At the same time the graduating or iris diaphragm must be brought into use.

Dr. Clifford Mercer, the President of the American Microscopical Society, who has quite recently reinvestigated the question of illumination, utterly condemns the narrow cone, as well as that of oblique light in all such investigations, and considers the 3-4ths axil cone as the most suitable method for microscopical illumination, and he bases his resolving limit accordingly. Some important experiments are brought forward by Dr. Mercer, which at the same time demonstrate the correctness of Lord Rayleigh’s limit of resolution (referred to in a previous chapter, p. 44), for circular apertures as contrasted with that calculated by the late Sir George Airy.

With regard to the Abbé Theory, Dr. Mercer says: “Resolution in the Abbé Theory may be said to increase by bounds. So long as the central image of the source of light alone is to be seen at the back of the objective, resolution is not present. The aperture may be increased without change in the contraction of the diffraction pattern, and in accompanying resolution, so long as the central image alone is to be seen at the back of the objective; but the moment the increase in aperture is sufficient to uncover or admit one flanking spectrum image, resolution is present. With greater increase in aperture, no improvement in the picture as to the contraction of the diffraction pattern is to be seen until another spectrum image is uncovered or admitted. Dr. Mercer gives his reasons for considering that the advantageous reduction in a cone of light between an object and the objective should not exceed, in the case of first-class objectives, one-fourth to one-third (never more than one-half) of the diameter of the cone. On the other hand, with full cone illumination, resolution increases continuously, and not by jumps or by periodic accessions. With regard to the use of oblique light, he says his Photos 2, 3, and 4[89] are a pictorial warning for a second time against the use of oblique illumination in ordinary work us a means of increasing, or of attempting to exhaust the resolving power of the microscope. At the same time it becomes evident that every substage should be provided with a means by which its condenser may be accurately centred, and that every student using the microscope should be familiar with a method of centring his substage condenser.

Dr. Mercer summarises the results of his experiments thus:—

1. “Diffraction rays on leaving an object may be considered in the same category with other rays changed in direction by an object.

2. “The diffraction phenomena seen in a projected image are essentially the effect of changes in light above the objective, due to a function of aperture, and not to changes below the objective, due to diffraction of light in the plane of the object.

3. “Diffraction in the plane of the object does, under some circumstances, furnish light to certain parts of an aperture from which primary rays are absent, and this enables aperture to more fully determine the character of the projected image, resulting in a more nearly truthful image, or, on the other hand, in false appearances. This is the gist of the Abbé phenomena of microscopic vision.

4. “But such phenomena are not peculiar to microscopic vision, notwithstanding Professor Abbé’s claim to the contrary.

5. “With any positive lens similar and more brilliant results may be got by utilising corresponding pencils of primary rays, instead of isolated pencils of diffracted rays.

6. “Still more trustworthy results may be got by using primary rays in place of the isolated pencils of primary rays.

7. “An advantage peculiar to using narrow cone illumination with an objective of wide aperture (the only illumination admissible in the Abbé theory), consists in giving, under suitable conditions, approximately the acme of resolving power simultaneously in each several diameters. Thus a circular aperture is approximately squared or made rectangular as to resolving power in several of its diameters simultaneously.

8. “Special attention is called to the fact that the Abbé theory deals with complex objects; for only such objects are subject to resolution. Single particles and uniform areas are outside its domain. These latter, however, are microscopic objects, and all objects are essentially different shaped aggregations of points. An isolated point-like particle, no matter what its minuteness, may be seen if it present sufficient contrast with the surrounding microscopic field. The size of the disc image is no less than a limit determined finally by aperture. That limit in size varying inversely with aperture, determines the limit of resolving power. This is the gist of the theory of microscopic vision which harmonises with our experimental study of aperture.”