5-9 THE YEAST CELL 



followed by Rafalko's Feulgen, Heidenhain, acid fuchsin -methyl 

 green, pyronln -methyl green have been studied. 



MATERIALS AND METHODS 



The yeast used was grown at room temperature on agar slants 

 in test tubes. When slides were to be made, a sample of yeast was 

 transferred from a 24 -hour slant growth to a flask of liquid medium 

 and incubated at 30°C. on a shaker for a period of from one to six 

 hours. After centrifuging a sample from the flask, cover-slip smears 

 were made without the use of albumin and were fixed by putting them 

 into a Petri dish of fixative. Throughout the staining process, the 

 material was handled in this way until mounted on a slide. At no time 

 was the material allowed to dry. 



The fixatives most commonly used were Navashin, Flemming, 

 Carnoy, Bouin, mercuric chloride and osmic acid vapors. Best re- 

 sults were obtained with Navashin and Carnoy. Staining procedures 

 employed included the Feulgen reaction, acid fuchsin -methyl green, 

 pyronin -methyl green, safranin-fast green, crystal violet-methyl 

 blue, Heidenhain's hematoxylin, Giemsa and Sudan IV. 



PROCEDURES AND SCHEDULES 



It might be well to note here that, with the possible exceptions 

 of the Feulgen reagents and Heidenhain's, it is best to employ fixa- 

 tives and stains that have been prepared immediately prior to their 

 use. This is especially true of fixatives, if best results are to be 

 obtained. In all the schedules given below, methyl alcohol and tolu- 

 ene were used in preference to ethyl alcohol and xylol because the 

 former are smaller molecules and therefore penetrate faster and 

 more evenly. 



1. Feulgen-fast green. The method of Rafalko (1945) was used. 

 This schedule is given on p. 5-8. 



Results: Chromosomes - red-red 

 Centrosome - purple -red 

 Nucleolus - pale green 

 Cytoplasm - pale green 



2. Acid fuchsin-methyl green. In separate bottles a 0.1% aqueous 

 solution of acid fuchsin and a 0.1% aqueous solution of methyl green. 

 When ready to use, two parts of the acid fuchsin were mixed with 

 4-5 parts of methyl green. The standard procedure calls for acidu- 

 lating every 50 ml. of the acid fuchsin with one drop of a 10% solu- 

 tion of acetic acid just prior to mixing the two dyes. Since that 

 made no difference in the staining results, that step was generally 

 omitted. 



