STAINING METHODS. 35 



for mounting i. e. , during the washing and dehydrating processes 

 usually employed. For staining, Kiihne prefers a methylene-blue 

 solution prepared as follows : Methylene blue, 1.5 parts; absolute 

 alcohol, ten parts ; triturate in a watch glass and add gradually one 

 hundred parts of a solution of carbolic acid containing five parts in 

 one hundred of water. The section is placed in this solution for about 

 half an hour, then washed in water and decolorized in a weak solution 

 of hydrochloric acid ten drops to five hundred grammes of water. 

 This part of the operation must be conducted very carefully, and 

 usually thin sections will only require to be dipped in the acid solu- 

 tion for an instant, after which they must be at once immersed in a 

 solution of lithium eight drops of a saturated solution of carbonate 

 of lithium in ten grammes of water. They are then allowed to re- 

 main in a bath of distilled water for a few minutes, after which they 

 are dipped into absolute alcohol, which Kiihne colors by the addition 

 of methylene blue. The sections are then placed in aniline oil which 

 contains a little methylene blue in solution, where they are dehy- 

 drated without the color being extracted from the stained bacteria 

 present. The aniline-oil blue solution is prepared by adding an ex- 

 cess of dry methylene blue to a small quantity of clarified aniline 

 oil. The undissolved pigment settles to the bottom, and a few drops 

 of the colored solution are added to a little aniline oil in a watch 

 glass to make the colored dehydrating bath. The section is next 

 washed out in pure aniline oil not colored after which every trace 

 of aniline oil is to be removed by the use of xylol. The section is 

 cleared up in turpentine and mounted in balsam. 



Ziehl-Neelson Method, for the tubercle bacillus in tissues. 

 Leave the sections for fifteen minutes in carbol-fuchsin solution 

 (No. 3) ; decolorize in sulphuric or nitric acid, twenty-five-per-cent 

 solution ; wash in sixty-per-cent alcohol ; place in a saturated aque- 

 ous solution of methylene blue for contrast stain ; wash, dehydrate, 

 and mount in balsam. 



The following method of staining sections for the purpose of 

 demonstrating bacteria present in the tissues is recommended by 

 Pregl (1891) as a substitute for the method of Kiihne. The results 

 are said to be excellent, and it is much simpler and more expeditious. 



The sections are made from tissues embedded in paraffin, and are 

 attached to clean glass slides with albumen-glycerin. Or they may 

 be attached to a cover glass by the following method when not em- 

 bedded in paraffin : The sections, completely dehydrated, are taken 

 out of absolute alcohol on a thin glass cover, upon which they are 

 extended ; a piece of filter paper is applied to the side of the cover 

 glass to absorb the alcohol, and before the section is completely dry 

 a drop of aceton-celloidin solution is placed upon it by means of a 



