WORKING APERTURE OF AN OBJECTIVE 327 



Mirror 



The substage concave mirror is usually so constructed as to deliver 

 light of about 0.25 N.A. Consequently, it will not fill the entire aper- 

 ture of objectives of higher power than 16 mm, of 0.25 N.A. (See Fig. 

 VIII-2.) 



Use of Diaphragm without Substage Condenser 



The purpose of the substage diaphragm is to modify the light and by 

 its aid obtain improved results in definition of the object which, with- 

 out it, are impossible. When the substage diaphragm is essentially 

 in the plane of the object, the limiting factors, so far as numerical aper- 

 ture is concerned, are the diameter and equivalent focal length of the 

 concave mirror. 



An opening in the diaphragm of about the same size as the front of the 

 objective is always used. The last use to which a substage diaphragm 

 should be put is to control the intensity of the illumination. The 

 intensity must be controlled with the aid of filters or dimmers. 



In the examination of most microscopic preparations, the problem is to 

 differentiate structure difficult to see, because its color or opacity 

 differs so little from its surroundings, rather than to observe detail at 

 the limit of resolution of the objective. The substage diaphragm is 

 found extremely helpful in examining such specimens by increasing 

 contrast and improving definition. Contrasts in the image can be 

 increased by increasing the size of the diffraction rings around the 

 image. Under these conditions the objective must be used at a lower 

 numerical aperture than that at which it is rated, which is accomplished 

 by closing the substage diaphragm. Care must be taken, however, so 

 as not to confuse diffraction patterns with true details of the specimen. 



If the substage diaphragm is closed so as to produce a reduction of 

 working aperture, the resolving power of the lens is lessened. However, 

 it is sometimes desirable to sacrifice some resolving power to improve 

 the contrast and visibility, especially when viewing specimens whose 

 index of refraction differs only slightly from that of the surrounding 

 medium. 



In actual practice, the simple substage diaphragm without the con- 

 denser can act as a field stop in partly reducing glare and extraneous 

 light, or it can be used to reduce the numerical aperture slightly. 



Working Aperture of an Objective 



The actual working aperture of an objective is equal to one half the 

 sum of the numerical aperture of the objective and the numerical 



