EMBRYOLOGY OF COCCIDS 79 



One of the fixing reagents most extensively used was Carnoy's 

 aceto-alcohol-chloroform mixture, prepared by mixing thoroughly 

 equal parts of absolute alcohol, glacial acetic, and chloroform 

 saturated with corrosive sublimate. Eggs fixed in this mixture 

 for from one to two hours lost their red pigment and became 

 transparent. They were then washed in 30 per cent alcohol for 

 two hours, and passed through 50 per cent to 70 per cent alcohol 

 with an intermission of one hour. They were then either left in 

 70 per cent alcohol until needed or dehydrated by passing them 

 up through 90 per cent, 100 per cent alcohol to x^dol and 

 imbedded in 52° to 58° paraffin. 



Sections were cut from on to 7m in thickness and stained mostly 

 with iron alum haematoxylin followed by eosin, orange G, acid 

 fuchsin, or a mixture of these. The triple stain, saffranin-gentian- 

 \dolet, and orange G, was also frequently used with very good 

 results. 



For whole mounts, the eggs were passed from 70 per cent to 30 

 per cent alcohol, in which the chorion was dissected away under 

 a binocular microscope. Embryos thus freed of their chorions 

 were stained with a diluted solution of gentian violet for from one 

 to six hours and then decolorized with 70 per cent alcohol. Dela- 

 field's haematoxylin, borax carmine, and alum cochineal have also 

 been used with fairly good results. The most beautiful specimens, 

 however, were those that were treated with gentian violet. 



For the study of the history of the germ cells not only the 

 genital organs of the embryo, but also those of several stages of 

 larvae, pupae, and adult scales were necessary. Ovaries were 

 mostly dissected out and fixed in either Fleming's or Zenker's 

 solution. In many cases, however, whole larvae, pupae, and 

 adult were put directly into Gilson's or Carnoy's aceto-alcohol- 

 chloroform solution, sectioned and stained in the same manner as 

 in the case of embryos. 



Perenyi's solution was also tried, but all except the aceto- 

 alcohol-chloroform mixtures w^ere useless unless heated to 70°C., 

 because the eggs as well as larvae are covered with a waxy sub- 

 stance which prevents penetration of fluids. AVhen heated, these 



JOUBNAl OF MOPa>HOLOGY, VOL. 33, NO. 1 



