286 STAINS AND STAINING. 



Kleine fib erg's Solution {Improved Formula). 



Haematoxylin, 2 J grammes ; crystallised calcium chloride, 

 20 grammes in 10 c.c. of distilled water; alum, 3 grammes in 

 16 c.c. of distilled water; rectified spirit, 240 c.c. Dissolve the 

 calcium chloride and the alum in their respective quantities of 

 water by the aid of heat ; mix the solutions and immediately dilute 

 with rectified spirit ; after an hour filter and add the haematoxyhn. 

 This makes a good working solution which keeps well. Of course, 

 it contains the alumina in solution not as alum, but aluminium 

 chloride. If in special cases the colour is considered too strong, 

 the dilution (when staining in bulk) must be made with some of 

 the solution to which haematoxylin has not been added. 



Ehrlich's H(E7natoxylin. 



This differs from the other two forms, in being strongly acidi- 

 fied with acetic acid. 



The usual book formula is haematoxylin, 2 grammes ; absolute 

 alcohol, 100 c.c; glycerine, 100 c.c. ; distilled water, 100 c.c; 

 glacial acetic acid, 10 c.c. ; "alum to saturation," 2 grammes being 

 the maximum quantity retained in solution at the ordinary tem- 

 peratures. 



It is generally admitted that when freshly made, Ehrlich's solu- 

 tion is next to useless, and we are told that it takes months, if not 

 years, to "ripen." The fact is that the rapidity depends upon 

 exposure. One method is to allow the solution to lie exposed in 

 an open dish till its volume is reduced to half, which gives it, of 

 course, the benefit of concentration as well as oxidation. 



It is, however, on absorption of oxygen that the whole question 

 of haematoxylin ripeness practically depends ; and, as this is 

 accelerated in alkaline and greatly retarded in acid solutions, it is 

 obviously more rational to effect the change upon the haematoxylin 

 before rather than after conversion into Ehrlich's solution. 



An " ammoniated haematoxylin " was therefore made by expos- 

 ing to the air a solution of haematoxylin in proof spirit made 

 alkaline with carbonate of ammonia, and afterwards converted 

 into Ehrlich's solution in the usual way. A staining fluid made on 

 these lines may be used at once and found superior to a normal 

 Ehrlich's solution which may have been ripening in the orthodox 



