ZOOLOGY AND BOTANY, MICROSCOPY, ETC. 
599 
acid, or in certain cases with sulphuric acid. It is then fixed with 
Kolossow’s mixture of uranium nitrate and osmic acid. Tho material 
may be preserved in thymol water (0 * 5 grm. to 1 litre). The sections 
are stained with a saturated aqueous solution of s ifranin or with anilin- 
oil-safranin, and, after having been washed with water are treated with 
a 2 per cent, solution of orange G, which dissolves the dye from the 
cell-wall and leaves the protoplasm and connecting threads stained. 
The threads may now be further stained by substitution, gentian-violet 
and Gram’s method giving the best results. After either the dye 
should be removed from the cell-wall by means of a 5 per cent, solution 
of acid fuchsin. After immersion in dilute glycerin, the sections are 
mounted in glycerin-jelly. 
(2) Iodine-acid violet method. The tissues are killel and fixed with 
iodo-potassic iodide solution, with an average strength of 0*5 per cent, 
iodine and 0*75 per cent, potassic iodide. The sections are caused to 
swell by immersion in sulphuric acid varying in strength according to 
the tissue from 1 to 30 per cent. The sections to bo stained are first 
mordanted with a solution of iodine in 5 per ceut. sulphuric acid. The 
staining solution is made by mixing equal parts of two stock solutions, 
the first being a 10 per cent, solution of II 2 S0 4 , and the second a 0*5- 
1 per cent, solution of pyoktanin or gentian-violet in water. 
To this staining solution the sections are directly transferred from 
iodo-potassic iodine and sulphuric acid mixture. The staining solution 
is allowed to act for 10 minutes, and then the sections are removed, and 
washed in water. The staining may be intensified by repeating the 
process. 
The sections should bo mounted in glycerin containing a small 
percentage of zinc chloride and a trace of iodine. 
The mounting medium is made by mixing 30 ccm. glycerin, 60 ccm. 
water, and 10 ccm. of a 20 per cent, solution of zinc chloride, adding 
a flake or two of iodine, and heating over a warm bath. 
Preparing Agar Media.* — Dr. M. P. Ravcnel gives the two follow- 
ing simple and rapid methods for preparing agar media, (i.) To make 
1 litre of agar, take (A) dried pepton (l per cent.), 10 grm.; salt (0*5 
per cent.), 5 grm. ; Liebig’s ext. (0*5 per cent.), 5 grm. ; water, 500 ccm. 
Boil for 3 minutes, and neutralise. (B) Agar (1*2 per cent.), 12 grm. ; 
water, 500 ccm. Chop the agar and put into autoclave, liun the auto- 
clave up to 2 atmospheres, giving 12 LI 0 heat. As soon as this pressure 
is reached, turn out the flame and allow the autoclave to cool down to 
below 100° before opening. The two solutions A and B are then mixed, 
cooled to 60°, the whites of two eggs beaten in 50 ccm. of water added, 
well stirred in, and the whole then boiled, and filtered through paper. 
Time, an hour and a quarter to an hour and a half. 
(ii.) To make permanently clear agar, fresh meat should be used as 
follows. To make 1 litre take (A) chopped meat, 500 grm. ; water, 500 
ccm. Mix, and place in a cool place over-night, then strain through towel. 
( B ) Agar (1 *2 per cent.), 12 grm. ; water, 500 ccm. Put in autoclave, run 
up to 2 atmospheres, put out flame, and cool down to below 100° before 
opening. When the agar solution has cooled down to about 75°, mix 
* Joum. Applied Microscopy, i. (1898) p. 106. 
