172 Animal Micrology 



stain varies from a few seconds or minutes for some of the dyes 

 (especially when used for cytoplasm) to 24 to 36 hours for others 

 (especially nuclear). Sections usually overstain, in which case 

 they are differentiated by means of alcohol, either pure or slightly 

 acidulated with hydrochloric acid. The color is thus extracted 

 rapidly; decolorization should be stopped immediately after the 

 color ceases to come from the tissue in clouds (20 seconds to 3 

 minutes). If acidulated alcohol is employed, it must be in much 

 weaker solution than that used for extracting carmines or hema- 

 toxylins. One part of hydrochloric acid to 1,000 of water or 

 alcohol is about the correct proportion. When one desires to 

 study the karyokinetic figures of nuclei, the acid-alcohol differen- 

 tiation should be employed, but if resting nuclei are to be studied, 

 only neutral alcohol should be used. 



30. Anilin Blue and Orange G (Mallory's connective tissue 

 stain). 



a) Double. 

 Solution I. 



Phosphomolybdic acid . 1.0 gram 



Distilled water . 100.0 c.c. 



Solution II. 



Anilin blue (soluble in water) 0.5 gram 



Orange G 2.0 grams 



Oxalic acid 2.0 grams 



The tissue should be fixed in mercuric chloride or in Zenker's 

 fluid. Sections are first placed in Solution I for one or two min- 

 utes and then washed well in water. They are then transferred to 

 Solution II for from 2 to 20 minutes, washed in water, dehydrated 

 rapidly, and cleared in xylol. This is an especially differential 

 stain for connective tissue fibrils which are colored a deep blue. 

 Keratin, blood corpuscles, and usually muscle and some nuclei, 

 are stained yellow; mucin and amyloid substances, a light blue. 



b) Triple. Stain the sections from 1 to 3 minutes in a 0.1 

 to 0.5 per cent, aqueous solution of acid fuchsin and wash them 

 in water before treating with Solution I as above. Further 

 procedure is the same as for a. 



