NEW METHOD FOR GRAM STAIN 177 



penetrated so deeply. The typhoid organism showed less pene- 

 tration of the Safranin than the staphylococci, in this respect 

 resembling Neisseria catarrhalis, but an occasional organism 

 showed a very definite violet center with a Safranin margin. 

 The Safranin masked the violet dye in the staphylococci more 

 rapidly than in the other two organisms. 



Experiment 5. The experiment was repeated with films fixed 

 by heat in the usual manner. The results were similar to that 

 obtained with the unfixed films. 



Experiment 6. The experiment was repeated with unfixed 

 films with the exception that the smears were partially decolor- 

 ized by a brief exposure to acetone and ether (equal parts) 

 before being stained with the Safranin. Upon examination 

 many of the typhoid organisms showed a very definite purple 

 center surrounded bj' a Safranin margin. Some of the typhoid 

 organisms showed irregularly spaced blackish granules in the 

 center somewhat resembling a string of cocci and surrounded 

 by a Safranin colored margin. Other of the typhoid organisms 

 appeared uniformly Safranin colored. As far as the eye could 

 determine the violet dye had penetrated to the center of many 

 if not all of the typhoid organisms. 



We conclude from these experiments that in ordinary Gram 

 staining the violet dye penetrates through the cell wall of Bacter- 

 ium tijphosum. Our experiments failed to demonstrate that' the 

 cell membrane of typhoid like organisms oilers much if any greater 

 resistance to the dyes than the cell wall of Neisseria catarrhalis. 

 The use of weaker dyes might bring out difference* not e\adent 

 when stronger dyes are used. The solution of methyl violet 

 used by us was of the same strength as used by Benians in one 

 of his experiments with Bacterium coli in which he apparently 

 demonstrated that the dye did not penetrate to the center of 

 the cell.' 



• With the Gram stain used by us the typlioid organisms decolorized slightly 

 more rapidly than the A^eisKeria catarrhalis organisms but as the latter were 

 distinctly larger the difference in rate of decolorization may have been due to 

 the difference in size. 



