Reagents 21 



sac of an angiosperm, or the free nuclear stage in the endosperm 

 of a gymnosperm, 2 per cent, or even 3 per cent, may be used; but 

 the finer details of the nucleus and cytoplasm are damaged by such 

 strong solutions. 



It will be found convenient to have in the laboratory the following 

 stock solution of chromo-acetic acid from which various solutions 

 can be made as they are needed : 



Chromic-acid crystals 10 g. 



Glacial acetic acid 10 c.c. 



Water 1,000 c.c. 



To make a solution containing 0.5 g. of chromic acid and 2 c.c. 

 of glacial acetic acid to 100 c.c. of water, add 50 c.c. of water to 50 c.c. 

 of the stock solution, and then add to the weakened solution 1.5 c.c. 

 of glacial acetic acid. Any desired proportions can be secured in a 

 similar way. Weighing the crystals for every new proportion is 

 more tedious. The proportions of the various ingredients, for the 

 present at least, must be determined by experiment. With favorable 

 objects like fern prothallia, Spirogyra, and other things which can 

 be watched while the fixing is taking place, suitable proportions are 

 rather easily determined, because specimens, after being placed in 

 the reagent, may be examined at frequent intervals, and combinations 

 which cause plasmolysis may be rejected and different proportions 

 tried until satisfactory results are secured. For example, fern pro- 

 thallia might be placed in the following solution: chromic acid, 2 g.; 

 acetic acid, 1 c.c.; and water, 97 c.c. If plasmolysis takes place, 

 weaken the chromic or strengthen the acetic, since the chromic 

 has a tendency to produce contraction, and the acetic to cause 

 swelling. A good fixing agent for fern prothallia can be made by 

 adding 50 c.c. of water and 1 c.c. of glacial acetic acid to 50 c.c. of 

 the stock solution. This solution will cause practically no plas- 

 molysis, and the fixing is thorough, but it must be remembered 

 that the proportion of acetic acid is rather high for cytological 

 details. A combination may be quite satisfactory for fern pro- 

 thallia and still fail to give good results with Spirogyra, and a com- 

 bination which succeeds very well with Spirogyra may not succeed 

 at all with Vaucheria. For very critical work the most favorable 



