GRAMS METHOD 101 



The bacillus of cholera, typhoid, glanders, the spirilla of 

 recurrent fever, the gonococcua and the Pneumococcus 

 Friedlanderi are amongst the organisms which yield up 

 their colour, and therefore do not stain by Gram's 

 method. 



The process is as follows: (1) Stain the cover-glass or 

 section in Ehrlich's aniline gentian violet for ten minutes. 

 (2) The section is then immersed without washing in iodine 

 solution (1 gramme of iodine and 2 grammes of potassium 

 iodide are dissolved in 300 c.c. of water) for one to two 

 minutes. (3) Wash in alcohol until no more colour comes 

 away. (4) Counter-stain in an aqueous solution of eosine. 

 (5) Dehydrate in alcohol. (6) Clear in xylol or oil of cloves, 

 transfer to the slide with the section-lifter, lay out flat, 

 blot off the excess of oil, add a drop of balsam, and mount. 



Gram-Giintlier Method. — Giinther has modified Gram's 

 original method by giving the preparation a washing with 

 a 3 per cent, solution of hydrochloric acid for a few seconds 

 after the first alcoholic washing. By this treatment cleaner 

 and brighter preparations are said to be obtained. Botkin 

 recommends that the section should be washed in aniline 

 water, after staining with gentian violet, and before im- 

 mersing in the iodine solution. 



It is very important to note that every pigment is not 

 suitable for this method. Fuchsine, methylene blue, and 

 Bismarck brown cannot be used, but only the so-called 

 pararosanilines, to which class belong methyl violet, gentian 

 violet, Victoria blue, etc., the strong affinity which these 

 colouring matters have for iodine being, according to 

 Unna, the cause of the remarkable action of Gram's 

 method. 



