43 



treating with l/10th per cent, acetic acid before staining. The stain is useful for 

 some cytological work (nuclear degenerations, cytoplasmic transformations, etc.). 

 In staining use the stock solution diluted with distilled water 1 :60 and rendered 

 slightly acid by the addition of 0.2% acetic acid, drop by drop, until the red color 

 tone, due to the fuchsin acid, becomes slightly accentuated. The success of the 

 stain depends upon having the staining reaction right; if too acid, the fuchsin 

 acid predominates, otherwise the green and orange prevail. Stain 6 hours or 

 more; when sufficient, dehydrate rapidly with absolute alcohol, clear with xylene, 

 mount in xylene balsam. 



112. Flemming's Triple Stain. Stain sections in an alcoholic solution of 

 safranin diluted with an equal volume of anilin water, for a day or longer. Differ- 

 entiate in absolute (or 95%) alcohol with l/10th % Hcl until hardly any more 

 color comes away. Stain for 1 to 3 hours in a 1% aqueous solution of gentian 

 violet. Rinse in distilled water and treat with a strong (2%) solution of Orange G. 

 in distilled water, and while clouds of violet are still being given off, bring the 

 sections into absolute alcohol in which the differentiation is begun. Transfer the 

 sections to clove oil or bergamot oil which completes the differentiation and clears. 

 Mount in balsam before the last pale clouds of color have ceased to come away. 



This stain is only recommended after such fixers as Flemming's fluid or Her- 

 mann's fluid. The stain is somewhat fickle, giving the best results only after some 

 practice. It is useful in some cytological work. 



Several modifications have been proposed. A short method is frequently 

 employed as follows: (1) stain for a second or two in a mixture of equal parts of 

 saturated aqueous and saturated alcoholic solutions of safranin; (2) rinse in 

 water; (3) stain 2 to 10 minutes in 1% gentian violet; (4) rinse in water; .and (5) 

 stain for 10 seconds or longer in a 2% solution of orange G.. Dehydrate rapidly 

 with absolute alcohol; clear and differentiate with clove oil, controlling the 

 differentiation under the microscope. Remove the clove oil with xylene or toluene 

 and mount in balsam. 



113. Congo Red. A well known indicator, red in neutral or alkaline 

 solutions, turning blue in the presence of free mineral and many organic acids, 

 not affected by acetic, lactic or carbonic acid in the presence of ammonia. It is 

 useful as a plasma stain after hematoxylin, gentian violet, etc. 



It may be employed in aqueous (^2 %) or alcoholic (2%) solution. With 

 subsequent differentiation in acid alcohol ( 110) it is a useful stain with gastric 

 glands. Occasionally useful as an indicator with living organisms or tissue. 



114. Eosin. Formulas: (a) %% aqueous solution; (b) 2-% aqueous 

 solution; (c) 1/10% solution in water or 95% alcohol. Formula (a) is preferable 

 for most work; (b) affords a stronger and (c) a weaker stain. This may be used 

 as a counter-stain with hematoxylin to differentiate nucleus from cell-body. 

 Stain sections after hematoxylin for 10 to 30 seconds, wash away the excess of 

 stain with distilled water or 67% alcohol. Since alcohol tends to wash out the 

 eosin, unless the color is too strong it is advisable to hasten the process of washing 

 out and dehydration. 



115. Erythrosin. Formulas: (a) l /z to 1% solution in 67% alcohol, 

 (b) ^4 to 1% aqueous solution. This is a general stain similar to eosin in its 



