76 BACTERIOLOGY 



Arens' chloroform method. — In order to avoid heating, aS 

 well as the preparation of a complicated staining iluid, an 

 alcoholic solution of fuchsine mixed with chloroform is used 

 as the dye, and alcohol with hydrochloric acid for decolorising. 

 The fuchsine solution is prepared by pouring 3 drops of 

 absolute alcohol on a crystal of fuchsine the size of a millet- 

 seed in a watch-glass and adding 2 to 3 c.cm. chloroform. 

 The solution becomes turbid and then begins to clear, 

 flocculent particles of fuchsine being separated. When 

 the clearing is complete the cover-glass preparation is laid 

 in it for from 4 to 6 minutes, until the chloroform is evapo- 

 rated, and is then decolorised in concentrated alcohol, to 

 which hydrochloric acid has been added in the proportion 

 of 3 drops to a watch-glass full. It is next rinsed in water, 

 and finally double stained with dilute methyl blue. 



Gram's decolorising method. — This appears to be the 

 most extensively used of all the bleaching methods, and 

 depends upon the employment of iodine in aqueous solu- 

 tion combined with potassium iodide (1 part iodine, 2 parts 

 potassium iodide, and 250 parts water) after the prepara- 

 tion has been stained in aniline water solution of gentian 

 violet. The iodine forms with the colouring matter a 

 precipitate which adheres to the micro-organisms, but can 

 be easily washed out of the tissues, and if this is properly 

 done the bacilli or cocci appear isolated by the stain. The 

 following is the mode of procedure in carrying out Gram's 

 method : — The prepared cover-glass or section containing 

 bacteria is warmed in aniline water gentian violet ; too 

 strong heating, however, has an injurious effect. The best 

 mode of warming the solution is to place it for 15 minutes 

 on the water-bath ; or to hold it over the flame for 1 minute, 

 let the watch-glass cool for 3 minutes, then heat it again for 

 a minute more and let it cool again for two or three, and so 

 on, until the process has been repeated four or five times. 



