608 Journal of Applied Microscopy. 



solution and afterward put in a solution of carminic acid, and hence are said to be 

 stained with carmine, but the order may be reversed and the alum used last, and 

 then that is the staining reagent. It is also clear that the tissue does not always 

 take the same color as the staining liquid, as for example, a solution of white 

 indigo will stain wood fibers indigo blue on contact with the air, or a solution 

 of haematein, which is brown, will give a violet stain. 



Logwood is a colorless preparation when fresh, but fermented or ripened it 

 becomes dark red-brown, or carmine-red since the hematoxylin present has been in 

 part changed to haematein. This was used at one time quite extensively on account 

 of its cheapness, but the tannin of the wood is unfavorable to good results. 



Hcematoxylin (CjgHj^Og) is the coloring principle of logwood, and is 

 extracted by the use of ether containing some amount of water. The crystals 

 when fresh are colorless, but owing to ready oxidation soon become violet tinted. 

 Solutions oxidize in the presence of alkalies and pass into brown hcKmatein or 

 over into higher oxidation products. It contains a weak acid and forms salts, 

 which take up acids readily and become hoemateates. To produce a stain, haem- 

 atoxylin alone is good for nothing ; a base must be present with it, in one of the 

 following ways : in objects already prepared ; in a mordant used before or after- 

 wards ; in the tissue at the same time as the stain. As bases those mostly 

 used are aluminum, chromium, iron, and copper, and the salts which rise from 

 these; haematoxylin never contains these. 



Hematoxylin Alone. — To stain plant cells, tissue is first fixed in picric acid 

 and well washed, some stain is left on the slide, and ammonia fumes poured 

 over. Ammonium-haematein is formed and the nuclei, especially, stained 

 violet. Just what base comes into this reaction is unknown, probably iron and 

 copper. Animal tissues stain but little in this solution. What takes place in 

 using this stain depends entirely upon the mixture or methods used. 



Hceinatoxylln with Chro7niu7ii. — Objects were first treated with potassium-bi- 

 chromate and later with the monochromate, since this holds in the tissues better. 

 After treatment for from twelve to fourteen hours in a one-third per cent, aque- 

 ous solution of haematoxylin, the sections were left equally long in a one-half per 

 cent, solution of potassium monochromate, and then passed up through alcohol 

 and xylol into paraffin. Weigert's modification is especially for the sheaths of 

 the' central nervous system. Material is hardened in Miiller's or Erlicki's fluids, 

 and sections, before the tissue has become green, are put from one to two hours 

 at 35° to 40° degrees C. in a three-fourths per cent, of ten per cent, alco- 

 holic haematoxylin, which stain must be several days old and contain haematein. 

 Then after the sections have become black, they are put into a mixture of borax 

 (four gr.); red potassium ferrocyanide (five gr.), and water (200 cc), and left 

 till the gray matter is yellow. A modification is made by treating the sections 

 with one-half per cent, chromic acid for a few minutes before putting them in the 

 stain. Other modifications differ only in treating the overstained sections with 

 potassium hyper-manganate (one-fourth per cent, solution), and oxalic acid and 

 potassium sulphite (1-200 water). This leaves the gray substance colorless, and 

 the white blue, and the sections may be further stained with carmine. 



Hczmatoxylin and Iron. — As with copper and chromium, the iron is in chem- 



