58 AIDS TO BACTERIOLOGY 



xylol is dropped on the section, and a cover-glass laid on 

 in such a way that the drop of balsam covers up the 

 section, and extends over the whole under-surface of the 

 cover-glass, as in the case of simple cover-glass prepara- 

 tions. The preparation is now ready for examination 

 with the oil-immersion lens. If the section is fixed to the 

 slide, as is the case with paraffin sections, all the manipu- 

 lations are carried out on the slide, the slide being flooded 

 with the stains and various reagents, or immersed in pots 

 containing them. 



Loffler's Method. The sections are stained in L6 flier's 

 methylene blue for from ten to sixty minutes, then rinsed 

 in distilled water and slightly decolorised by immersing 

 in a 0-5 per cent, solution of acetic acid for a few seconds. 

 The sections are now washed, dehydrated in absolute 

 alcohol, cleared in xylol, transferred to the slip, blotted, 

 and mounted as usual. Carbol-thionine blue may also be 

 used, and often differentiates better than Loffler's 

 blue. 



Kuhne's Method. The sections are placed in Kiihne's 

 carbol methylene blue for thirty minutes or longer, then 

 washed in water, and very carefully decolorised in very 

 dilute hydrochloric acid (20 drops of strong acid in 100 c.c. 

 water). Thin sections only require to be immersed for 

 two or three seconds. The sections are at once trans- 

 ferred to an alkaline solution (10 drops of a saturated 

 solution of lithium carbonate in 10 c.c. of water), washed 

 in water for a few minutes, dehydrated in absolute alcohol 

 tinted with methylene blue. The sections are now placed 

 in anilin oil, which also contains a little dissolved methy- 

 lene blue. After being washed in colourless anilin, then 

 in xylol, they are mounted, as usual, in balsam. 



Ziehl-N eelsen Method. Where sputum is to be examined, 

 a little of the material is spread on a microscope slide. 

 If any small yellow caseous particles are present, these 

 should be selected, otherwise the thick portion of the 

 sputum should be used. After spreading, drying, and 

 fixing, filtered carbol -fuchsin solution is dropped on the 

 film, and the slide is heated over a Bunsen flame till 

 steam just rises, care being taken to prevent the stain 

 boiling. The stain is allowed to act for five minutes, 

 fresh stain being added as evaporation takes place. The 

 preparation is now decolorised by dipping alternately 



