ALIGNMENT OF THE PHASE MICROSCOPE 109 



2. Focus the microscope on the specimen. The image seen at first may be a poor 

 one. If the optical properties of the specimen are such th:it its image is detected 

 with difficulty with an ordinary nrici-oscope, it may become necessary approximately 

 to center the opening in the condenser diaphragm with respect to the conjugate area 

 of the diffraction plate in order to detect any part of the specimen. This is done by 

 replacing the eyepiece with the centering telescope, focusing the telescope on the 

 coating of the diffraction plate, and mani[)ulating the centering adjustments of the 

 condenser diaphragm until the image of the opening in the condenser diaphragm 

 appears over the conjugate area of the diffraction plate. Replace the eyepiece 

 and then focus the microscope on the specimen. If the substage condenser to- 

 gether with the diaphragm has been racked too far away from the specimen slide, 

 the central opaque area of the diaphragm may block too much of the light from 

 the lamp, and the center of the field of the microscope will appear dark. In fact, 

 if the substage condenser together with the diaphragm is either too far from or too 

 close to the specimen slide the center of the field of the microscope may appear 

 dark or colored or both, particularly if the iris diaphragm of the lamp has been 

 closed to a small aperture. Spherical aberration of the substage condenser may in- 

 troduce additional vignetting of light from the lamp, and chromatic aberration of 

 the substage condenser may introduce color in the field of the microscope. It is 

 sometimes advantageous to use a low-power phase objective as a finder. In 

 this connection, if a phase objective is used together with the condenser diaphragm 

 for a higher power phase objective, the center stop of the condenser diaphragm may 

 be large enough to simulate the conditions for darkfield illumination, and this can 

 also be helpful in locating some specimens. Before going to step 3, make certain 

 that the diaphragm in the substage condenser is the diaphragm supplied for the 

 particular phase objective. 



3. After the microscope has been focused on the specimen, close the iris diaphragm 

 on the lamp housing to form a small aperture. Raise or lower the substage condenser 

 until the image of the lamp diaphragm is focused on the specimen. If the substage 

 condenser is of the Abl)e type, the image of the edge of the lamp iris appears colored, 

 and more strongly so when the condenser diaphragm with an ainiular opening is in 

 position than when the condenser is used with a central clear aperture. The sub- 

 stage condenser is considered to be in good focus when the image of the edge of the 

 lani]) diaphragm is in the color range blue-green to purple-blue-purple. On account 

 of the spherical aberration of an Al)be-type sulistage condenser, the position of the 

 so-called good focus for this condenser will be different with annular than with 

 central openings in the condenser diaphragm. It may also be advantageous to 

 refocus this substage condenser when a phase objective which requires an opening of 

 one mean diameter and width in the condenser diaphragm is replaced by a phase 

 objective which requires an opening of another mean diameter and width. Adjust 

 the substage mirror so that the imag(> of the lamp diaphragm is at least approximately 

 centered in the field of view. 



4. Image the lamp filament on the condenser diaphragm or on the iris diaphragm 

 of the substage condenser by focusing the lamp condenser. If a plate of diffusing 

 glass has been placed in front of the lamp, it should be removed during this step, 

 but it may be replaced after the lamp has been adjusted. Full use should be made 

 of whatever means have been built into the lamp housing to adjust the lamp for 

 optimum illumination. The image of the filament should extend at least over the 

 annular opening in the condenser diaphi'agm. If the lamp envelope is frosted, the 

 envelope is imaged on the condenser diaphragm. 



