specialized Microscopy 97 



is then slowly raised until all oroanic matter is burned away and only 

 the mineral skeleton of the tissue is left. This appears white under 

 dark-field observation. 



Phase and Interference Microscopy 



Although most living material is transparent to visible light, different 

 components of tissue do alter to a different extent the phase of light 

 waves passing through them. That is, the light velocity is altered, 

 advanced or retarded and its vibration is said to be changed in phase. 

 When two waves come together and are in phase, brightness increases, 

 but if out of phase and of equal amplitude, interference occurs and the 

 eye sees black, not light. W'aves of intermediate difference in phase pro- 

 duce a series of grays. It is the aim of the phase microscope, to change 

 slight phase differences into amplitude differences and produce a vari- 

 ation in intensity from light to dark contrast observable in the specimen 

 tuider the microscope. 



Both phase and interference microscopes follow this principle of 

 interference phenomenon — combining light waves that are out of phase 

 Avith each other to produce combined ^vaves of greater and lesser 

 amplitude. The means of applying the principle is different. 



PHASE MICROSCOPY 



The cost of phase microscopy is not excessive and the parts can be 

 adapted to any brightfield microscope. The system requires phase con- 

 trast objectives (achromatic objectives Avith fixed-phase plates and a 

 t^vo-lens Abbe condenser with an iris diaphragm and a rexolving ring 

 below the condenser to carry four annular ring diaphragms (stops that 

 produce different-sized cones of light). The correct annular ring dia- 

 phragm must be centered to the phase ring of the correct objective and 

 match its numerical aperture. The anniUar diaphragm causes the light 

 to strike the object in the shape of a hollow cone and gives rise to two 

 types of waves; one type passes straight through the object (undiffracted), 

 the others are diffracted into a different course. Different components 

 in the tissue diffract differently producing the differences in intensity. 



Images under phase microscopy exhibit a "halo" as a result of the 

 diffraction of light at the phase-changing annulus (annular ring). The 

 light which has passed straight through the specimen is made to inter- 

 fere with the light diffracted sideways by it. Only the refracting struc- 

 tures are observed, and these edges and abrupt changes of refracti\e 

 index produce the "halo" aroinid the images. 



