656 GENERAL STAINING 



3 per cent, alum. Stain in safranin for twelve to fifteen hours. 

 Differentiate in weakly acidulated 70 per cent, alcohol until the 

 cytoplasm loses its rose tint. 



1312. Safranin and Crystal (or Gentian) Violet (Hermann, 

 Arch. mikr. Anat., xxxiv, p. 58), Stain three to twenty-four hours 

 in safranin (1 per cent, in 50 per cent, alcohol, made with anilin 

 water) ; destain five to thirty seconds in 50 per cent, alcohol. 

 Counterstain in a solution of crystal violet in anilin water for 

 ten seconds to a few minutes. Dehydrate rapidly through 50, 70 

 and 95 per cent, alcohols, omitting 50 and 70 per cent, if stain 

 lost too rapidly. Clear and differentiate in clove oil ; wash in 

 several changes of xylol. 



Chromosomes and nucleoli red ; resting nuclei and chromatin 

 granules violet ; prophase and early teloj^hase nuclei may show 

 violet chromonema and red chromatin granules ; cytoplasm light 

 violet and spindle deeper violet. 



Substitution of dilute solution of crystal violet in clove oil for 

 aqueous solution gives brilliant staining of nucleoli and chromo- 

 somes. Remove excess stain with clove oil. 



The staining may be reversed in effect. Stain six to twenty-four 

 hours in a 1 per cent, solution of crystal violet in 70 per cent, 

 alcohol and destain briefly with 50 per cent, alcohol. Counter- 

 stain with safranin for fifteen seconds to five minutes. Dehydrate 

 rapidly, clear in clove oil and wash in xylol. Chromosomes and 

 nucleoli violet, cytoplasm and spindle red. See also Stockw^ell 

 {Science, Ixxx, 1934, p. 121) for a safranin, gentian violet staining 

 method for difficult cases. 



Flemming's Triple Stain. See §§ 322 and 323. 



1313. Nebel's Crystal Violet and Acid Fuchsin {Zeits. Zellf. 

 Mikr. Anat., xvi, 1932, p, 251), Stain from water in acid fuchsin 

 and differentiate in picric acid. Stain in crystal violet and dehy- 

 drate rapidly, 



1314. Methyl Green and Acid Fuchsin (Guignard, Rev. Gen, 

 Bot., i, p, 11), Stain several hours in aqueous methyl green and 

 differentiate in water or 50 per cent, alcohol until the stain is 

 removed except from the nuclei and chromosomes. Stain for a 

 short time (thirty seconds to three minutes) in aqueous acid 

 fuchsin which tends to replace the methyl green. Dehydrate with 

 absolute alcohol, clear in clove oil, wash in xylol and mount. 

 Chromosomes and nucleoli green, spindle and cytoplasm pink. 



1315. Cyanin and Erythrosin. Stain a few minutes to an hour 

 or more in solution of cyanin in 70 per cent, alcohol. Rinse in 

 70 per cent, alcohol and stain a few seconds in erythrosin solution 

 in 70 per cent, alcohol. Dehydrate rapidly with 95 per cent, and 

 absolute alcohol. Clear in clove oil, wash in xylol and mount. 

 Chromosomes blue, cytoplasm pink. If stains lost too readily 

 omit 95 per cent, alcohol and clove oil. 



