MORPHOLOGY OF SYMBIOTIC BACTERIA 473 



The staining solutions are: 



1. Altmann's acid fuchsin anilin solution: 



Acid fuchsin 20 grams 



Anilin water 100 cc. 



2. 1 per cent solution of methyl green 



The sections after being prepared for the staining process by treat- 

 ment with permanganate of potassium followed by oxalic acid, are 

 stained for five minutes in the acid fuchsin solution which has been 

 previously warmed to 60°C. Next they are thoroughly washed in 

 distilled water, and dipped for an instant into the solution of methyl 

 green, then washed, rapid!}' dehj'dratcd in absolute alcohol (alcohol 

 of intermediate strength must be avoided) cleared in toluol, and mounted 

 in balsam. 



The author has found that the permanganate and oxalic acid 

 treatment may be omitted if the fixation has not been carried too 

 far. However, if the staining differentiation is not sharp the 

 sections may be treated for a minute or so in 1 per cent perman- 

 ganate of potassium and followed by a similar treatment in 5 per 

 cent oxalic acid. I have also found a great variation in the 

 quality of various brands of methyl green. I have only been 

 able to get good results with Griibler's methyl green. This 

 method, when carried out successfully, gives excellent differentia- 

 tion between bacteria and tissues. 



There are several mitochondrial methods in use. Benda's 

 crystal violet method, perhaps, gives the sharpest differentiation 

 of all these methods. This is a very long and tedious method, 

 but in some cases the results appear to justifj' the longer pro- 

 cedure. Other methods are: Altmann's anilin fuchsin picric 

 acid method, Schridde's modification of Altmann's method, 

 Regaud's method, the copper and iron hemotoxylin methods, 

 etc. I have stained bacteria by all these methods with good 

 results. 



The mitochondrial staining methods were devised to fix and 

 stain delicate bodies (mitochondria) in the cytoplasm which are 

 not visible after ordinary histological technique. It appears that 

 some symbiotic bacteria, particularly those that have an intra- 

 cellular relationship, are just as fragile and delicate as mito- 



