364 OSCAR W. RICHARDS 



other acid-fast bacteria {31, p. 452), and for detecting important 

 chemicals like riboflavin and vitamin A. Some observers believe 

 malignant tissues may be identified with the fluorescence microscope 

 (32). 



Since long wavelength ultraviolet is transmitted by optical glass 

 down to about 310 mju an ordinary microscope may be used. For 

 work with shorter wavelength radiation a quartz or Corex condenser 

 is required. The carbon arc is the strongest ultraviolet source, al- 

 though for many studies the mercury arc or the tungsten lamp may be 

 used. Filters are used to remove the visible light. 



Two methods are used. One method employs a darkfield condenser 

 so that none of the ultraviolet can pass directly into the objective. 

 The resulting fluorescence usually is less bright; the observer's eye 

 must be well dark-adapted and the observations made in a darkened 

 room. Photographic exposures may require hours. 



The other method uses the crossed filter technique. A filter that 

 transmits only the radiation to be absorbed {e.g., blue or ultraviolet) 

 is placed over the source; the eyepiece filter {e.g., yellow filter) does 

 not transmit this direct radiation from the source, but will let through 

 the fluorescent light. This procedure gives brighter images and 

 makes possible shorter exposures. When the fluorescence involves 

 all colors, the filter on the source must not transmit any visual light. 

 Since some ultraviolet will pass through the microscope, an ultra- 

 violet-absorbing filter should be used to protect the eye and is required 

 when photographs are made because many emulsions are more sensi- 

 tive to short wavelength radiation. The lens of the eye fluoresces 

 and obscures vision when such a protective filter is not used. Fluo- 

 rite fluoresces so that apochromatic and fluorite objectives are less 

 satisfactory than achromatic objectives. The mounting medium 

 and the immersion oil used must be nonfluorescing, which disqualifies 

 cedar oil, balsam, and many of the usual materials that fluoresce 

 strongly. Glycerin is a good mounting medium and some of the 

 modern synthetic immersion oils are suitable. 



The need for a darkened room and dark-adapted eyes restricts 

 fluorescence microscopy and the observations may be time-consum- 

 ing, since some of the phenomena are transient. The image is not 

 too bright and longer photographic exposures are required. The 

 method offers a promising field for the investigator desiring to work 

 in a realm with few known rules and guideposts and relatively un- 

 known possibilities {31-33). 



