LABORATORY STUDIES AND DEMONSTRATIONS. 219 



found by examining sweet cider microscopically. For the fol- 

 lowing methods of demonstrating nuclei in yeast and obtaining 

 ascospores we are indebted to Mr. S. C. Keith Jr. 



To Demonstrate Nuclei in Yeast. Any good actively-growing 

 yeast will answer, but a large (brewers') yeast is preferable. Mix 

 a little of the yeast with an equal amount of tap- water in a test- 

 tube and shake thoroughly. Add an equal volume of Hermann's 

 fluid and shake again. As soon as the yeast has settled pour off 

 the supernatant liquid and wash the yeast by decantation. Trans- 

 fer some of the cells to a slide, fix by drying, stain by Heiden- 

 hain's iron-hamiatoxylin method (see Centmlblatt far Bacteri- 

 ologie, xiv. (1893), pp. 358-360), wash, dehydrate with alcohol, 

 follow with cedar-oil, and mount in balsam. In successful speci- 

 mens the effect is very satisfactory. (See Fig. 96.) 



A Simpler Method. To demonstrate nticlei in yeast more 

 quickly and very easily the following method may be used : Boil 

 (in a test-tube) for a moment an infusion of very vigorous yeast 

 in water, place a drop of the boiled infusion on a slide, add a 

 drop of very dilute "Dahlia" solution, cover, and after one or 

 two minutes examine with a high power. The nuclei in most of 

 the cells will be easily discoverable. 



To Obtain Ascospores in Yeast. It has been usually recom- 

 mended to employ for this purpose blocks of plaster-of-Paris. 

 We have found the following method more trustworthy : 



The yeast to be used should be the " top" yeast used in ale- 

 breweries. It should also be actively growing and fresh. If 

 fresh yeast cannot be obtained, some may be revived by cultiva- 

 tion for 24 hours at 25 C. in wort, and a little of the thick sedi- 

 mentary portion may then be placed in a very thin layer on dry 

 filter-paper which has previously been sterilized by baking. The 

 filter-paper is then placed on a layer of cotton about inch in 

 thickness lying on a plate or saucer, the cotton having previously 

 been thoroughly wetted with cold sterilized tap-water. The 

 whole is covered by a bell-glass and set in a rather warm place 

 (25 C.). In the course of two or three days spores will be found 

 in many of the cells. The lower the temperature the longer is 

 the time required for spore formation. If "bottom" yeast is 

 used instead of "top" yeast a much longer time is required, and 

 the results are far more uncertain. 



