no MICROSCOPIC METHODS. 



5. Carbol-Fuchsin (see p. 1 13). This is a very powerful stain, and, 

 when used in the undiluted condition, ^-i minute's staining is usually 

 sufficient. It is better, however, to dilute with five to ten times its 

 volume of water and stain for a few minutes. In this form it has 

 a very wide application. Methylated spirit with or without a few 

 drops of acetic acid is the most convenient decolorising agent. Then 

 dehydrate thoroughly, clear, and mount. 



Various other staining combinations might be given, 

 but the above are the best and most widely used. If 

 the reader has thoroughly grasped the remarks made 

 above on the general principles which underlie the stain- 

 ing of bacteria, he will be able to use any combination 

 to which his attention may be directed. We may only 

 add here that different organisms take up and hold 

 different stains with different degrees of intensity, and 

 thus duration of staining and degree of decolorisation 

 must be varied. It may be laid down as a general rule 

 that, so long as organisms retain the stain, the greater the 

 decolorisation of the tissues in which they lie, the clearer 

 will be the results. 



Gram's Method and its Modifications. In the methods 

 already described the tissues, and more especially the 

 nuclei, usually retain some stain when decolorisation has 

 reached the point to which it can safely go without the 

 bacteria themselves being affected. In the method of 

 Gram, now to be detailed, this does not occur, for the 

 stain can here be removed completely from the ordinary 

 tissues, and left only in the bacteria. All kinds of bac- 

 teria, however, do not retain the stain in this method, 

 and therefore in the systematic description of any species 

 it is customary to state whether it is, or is not, stained by 

 Gram's method by this is meant, as will be understood 

 from what has been said, whether the particular organism 

 retains the- colour after the latter has been completely 

 removed from the tissues. It must, however, be remarked 

 that some tissue elements may retain the stain as firmly as 

 any bacteria, e.g., keratinised epithelium, calcified particles, 

 the granules of mast cells, and sometimes altered red blood 

 corpuscles, etc. 



