STKONG : DEVELOPMENT OF COLOR IN DEFINITIVE FEATHER. 149 



Material was kept in the picro-sulphuric solution for about five hours 

 and then transferred to 70% alcohol followed by 90%. It usuallj^ took 

 one to two weeks with several changes of alcohol to remove all traces 

 of picric acid. A fixation of three hours was found sufficient for 

 Hermann's fluid and the usual methods of washing and hardening 

 followed. 



Dehydration was accomplished by immersion in absolute alcohol for 

 at least twenty-four hours. 



For clearing and infiltration with paraffin, I have found the chloro- 

 form method especially satisfactory ; it was the only successful medium 

 for coruified portions of the feather when anything like complete series 

 w^ere desired. I have found it particularly good in preparing material 

 for sections of dry feathers. I have often secured almost perfectly 

 complete sei'ies with it, whereas with xylol, or cedar oil, only occasion- 

 ally would a section i-emain in the paraffin I'ibbon. 



Feather germs were left in melted paraffiii two to five days and were 

 then imbedded in hard paraffin (135° F.).^ Dry feathers were, in 

 ordinary cases, dropped into chloroform for a few hours and then 

 transferred to melted paraffin for about twelve hours. 



Serial sections were cut with a Minot-Zimmermann microtome 3^ to . 

 10 micra thick, mostly 3^ or 6§ micra. Also a few sections at the proximal 

 end of the feather germ were cut 2 micra thick by means of the Minot 

 microtome having Zimmermann's improved feeding attachment. I 

 found it necessary to have the temperature as low as 60° F., and each 

 section was cut with a very slow motion of the object carrier. For 

 almost all purposes, however, sections 3-^ micra thick are thin enough. 



Sections of the cornified portions of the feather germ are very elastic 

 and tend to curl and spring from the paraffin ribbon, especially when 

 the sections are as much as ten micra thick, but with the methods 

 described above fairly complete series -were obtained. 



Mayer's albumen fixative was used successfully for affixing sections to 

 the slide ; but with osmic-acid material it was found necessary to 

 spread, in addition, a thin film of celloidin over the sections, immediately 

 after the immersion in alcohol which followed the removal of paraffin 

 with xylol. This celloidin film held the sections securely in position and 

 did not interfere with subsequent work. 



A number of stains were tried, but l)y far the most satisfactory 

 were (1) for material fixed in picro-sulphuric a double stain, viz. 



^ A mixture of hard paraffin with about 5% of resin was suggested by Professor 

 G. H. Parker and was used with some success for dry feathers. 



