GRAM'S METHOD 87 



centrated alcoholic solution of methyl blue, twenty parts of a 

 ■^ per cent, eosine solution in 70 per cent, alcohol, and forty 

 parts water, and after staining are rinsed in water, and the 

 remainder of the treatment carried out in the usual way. 

 Plehn recommends the addition of twelve drops of a 20 per 

 cent, caustic potash solution to the water. 



Gram's metliod. — This method is in a high degree suited 

 for sections. They are stained in aniline water gentian 

 violet, to the action of which they are exposed for from ten to 

 thirty minutes ; but the time of staining may be shortened 

 by heating. After staining they are rinsed in water and 

 immersed for two to three minutes in a solution of iodine and 

 potassium iodide, and are then kept moving to and fro in 

 90 per cent, alcohol until no more colouring matter comes 

 away. The sections, which now appear of a slate-grey 

 colour, are next transferred to alcohol, cedar oil, and 

 Canada balsam. The bacteria are seen in violet on a 

 yellowish ground. Double-staining with picrocarmine or 

 Magdala red causes the violet tint of the micro-organisms to 

 stand out distinctly against the red colour of the tissue. 



The method of Gram may also be reversed, and the 

 sections first stained for fifteen minutes in picrocarmine or 

 Magdala red, rinsed in 50 per cent, alcohol, and then laid 

 in aniline water gentian violet. After decolorising in 

 iodine solution, the preparation is treated with alcohol, oil, 

 and Canada balsam. 



Giinther's modification, which is characterised by the 

 exposure of the sections, after decolorising in alcohol, to 

 the action of a 3 per cent, solution of hydrochloric acid, 

 yields brilliant results (compare p. 54). 



Kiihne's modification of Gram's process. — Gram's method 

 has undergone many further modifications in its use for 

 sections, and Kiihne in particular has devised a number of 

 processes, of which the following are the most important. 



