54 AIDS TO BACTERIOLOGY 



But the clear haloes sometimes seen round bacteria in 

 albuminous material must not be confused with capsules. 

 Bum's India ink method and Gram's method often show 

 the capsules very well. 



Richard Muir's Later Method. A thin film after drying 

 is stained with warm carbol-fuchsin for 30 seconds, 

 washed slightly in alcohol and again thoroughly in water. 

 It is placed in the following mordant for a few seconds: 



Sat. sol. mercuric chloride . . . . . . 2 parts 



Tannic acid (20 per cent.) solution . . . . 2 



Sat. sol. potash alum . . . . . . . . 5 



After well washing in water, the preparation is treated 

 with methylated spirit for about a minute and should 

 then be pale red. After thorough washing in water 

 and counter-staining with methylene blue for 30 seconds, 

 the film may be dehydrated in alcohol, cleared in xylol 

 and mounted, or may be simply dried with filter paper. 

 The bacteria are of a deep crimson, and the capsules blue. 



The Treatment and Staining of Sections. The tissue is 

 first ' hardened,' and then cut into sections with some 

 form of microtome. 



Hardening of Tissues. The most satisfactory harden- 

 ing reagent is alcohol. It is preferable to pass through 

 increasing strengths of alcohol e.g., 50 per cent., 75 per 

 cent., and finally absolute the tissue remaining in each 

 for twenty-four to forty- eight hours. The tissue is cut 

 into pieces 10 to 20 millimetres square and 5 to 10 milli- 

 metres thick; these are immersed in the alcohol, which 

 may be changed once. If absolute alcohol alone be used, 

 it may be contained in a wide-mouthed bottle, in the cork 

 of which are fixed several needles. The pieces of tissue 

 are placed on the needles in such a manner that, when 

 the cork is fixed in the mouth of the bottle, the pieces of 

 tissue are just beneath the surface of the alcohol. The 

 alcohol gradually abstracts the water from the tissue, and 

 as that containing the water sinks to the bottom, fresh 

 alcohol constantly comes in contact with the material. 

 Tissues containing much water are naturally more diffi- 

 cult to harden than those containing little. 



Before hardening in alcohol, it is usual to ' fix ' the 

 tissues, for the better preservation of the tissue element 

 A saturated solution of corrosive sublimate in water, to 



