STAINS 197 



GENERAL STAINING METHODS 



180. Methods of staining. There are two principal methods of staining, — 

 (1) in bulk or loose sections, and (2) on the slide. The second method gener- 

 ally follows the process of sectioning in paraffin, and is given special con- 

 sideration in Sees. 197-199. Staining in bulk is, on the whole, much less 

 precise in its results than the staining of microtome sections. The methods 

 of staining in bulk also apply to sections cut free-hand (Sec. 194), either 

 from preserved or living material. 



181. Eosin. Saturated solutions in water or alcohol are used. Material is 

 stained almost at once, and should then be transferred to 1% acetic acid for 

 a minute (which renders the stain less soluble), after which the acid should 

 be thoroughly washed out. Permanent preparations are generally made in 

 glycerin after the method outlined in Sec. 188. If the preparations are to 

 be mounted in balsam (Sec. 187), the staining should be with alcoholic solu- 

 tions, or, better still, with a solution in absolute alcohol from which the 

 material may pass directly into xylol, and there is no need of treatment 

 with acetic acid. 



Eosin does not stain cell walls and never overstains protoplasm. It is 

 especially useful for the fungi, which are generally mounted in glycerin, 

 and is one of the best of the quick, simple stains. 



182. Iron-alum haematoxylin. This method, developed by Heidenhain, 

 gives the most satisfactory results of all the hcematoxylin stains in the differ- 

 entiation of protoplasmic structure. Delafield's haematoxylin (Sec. 183) is a 

 somewhat better stain for tissues, because it colors cell walls sharply. Iron- 

 alum hsematoxylin does not color cell walls heavily, and is consequently a 

 very useful general stain for the algse and fungi which are to be stained in 

 bulk and mounted without sectioning. 



Two separate solutions are used : 



(1) A 2% aqueous solution of ammonia sulphate of iron (iron alum). 



(2) A i% solution of hsematoxylin dissolved in hot distilled water. 



The solution of iron alum acting as a mordant prepares the tissue to take 

 up the hsematoxylin. Bring the material from water (running it down 

 through the grades of alcohol, if preserved in the latter) into the iron-alum 

 solution, which for delicate structures may be diluted to l';^. Leave in the 

 iron alum from one to three hours, rinse for a few minutes in water, and 

 place in the hsematoxylin solution. If the hiematoxylin becomes too muddy, 

 replace it with fresh. Leave the material in the haematoxylin from three to 

 ten hours (over night does no harm) and then place in iron alum again. The 

 black stain extracts rapidly, and the material must be examined from time 

 to time under the microscope. When the stain has been extracted to the 

 proper point, place the material in considerable tap water for a half hour, or 



