Stains and Staining 47 



If, after rinsing in water, the stain is evidently too weak, put 

 the slide or section back into the stain until it appears overstaincd. 

 Place the slide in acid alcohol. If an acid alcohol with 2 drops of 

 HC1 to 100 c.c. of 70 per cent alcohol reduces the stain too much in 4 

 or 5 seconds, use less acid. Transfer to 70 per cent alcohol without 

 any acid. As soon as the color changes from red to purple, examine 

 under the microscope. If it is still overstained, return to the acid 

 alcohol; if the stain is too weak, return to the haematoxylin and 

 try it again. After the haematoxylin is just right, apply a contrast 

 stain, if you wish to double stain. Before transferring to the xylol 

 wipe the alcohol from the back of the slide, or at least rest the corner 

 of the slide upon blotting-paper for two or three seconds, in order 

 that you may not carry over so much alcohol into the xylol. Add 

 a drop of balsam and a cover. Since the xylol is very volatile, this 

 last step must be taken quickly. If blackish spots appear they 

 are usually caused by the drying of sections before the balsam and 

 cover are added; if there are whitish spots or an emulsion-like 

 appearance, the clearing is not thorough; this may be caused by 

 poor xylol (or other clearing agent) ; by absolute alcohol which is 

 considerably weaker than its name implies (the absolute alcohol 

 must test at least as high as 99 per cent, and ought to test as high 

 as 99.5 per cent, if xylol is to be used for clearing); or by passing 

 too quickly through the absolute alcohol and xylol, or even by 

 moisture on the cover-glass. The last danger is easily avoided by 

 passing the cover quickly through a Bunsen or alcohol flame before 

 laying it on the balsam. 



Delafield's haematoxylin is the most generally useful stain in the 

 haematoxylin group. It brings out cellulose walls very sharply, 

 and consequently is a good stain for embryos and the fundamental 

 tissue system in general. With safranin it forms a good combination 

 for the vascular system, the safranin giving the lignified elements a 

 bright red color, while the haematoxjdin stains the cellulose a rich 

 purple. It is a good stain for chromatin, and the achromatic struc- 

 tures show up fairly well, but can be brought out much better by 

 special methods. Archesporial cells and sporogenous tissue are 

 very well defined if proper care be taken. Lignified and suberized 



