IRON-HEMATOXYLIN-STAINED TESTIS 121 



Mayer's albumen, which may interfere with differentiation or staining, is used. 



The dry sections attached to the slides should be warmed on the underside 

 until the paraffin melts, placed in xylene until the paraffin is removed com- 

 pletely, and then run down in the ordinary way through absolute alcohol and 

 lower percentage alcohols to distilled water. Then they should be lifted from 

 the distilled water and examined carefiilly. If there is a tendency for the water 

 to gather in droplets on the slide or, if upon shaking the water from the slide, 

 each section appears to retain an adherent coat of water around itself, it is an 

 indication that either the wax was not removed properly in the xylene or 

 that the xylene itself is so old as to have a wax content too high to be useful. 

 Such slides must be returned through the alcohols to absolute alcohol and 

 thence to clean xylene where they should be left for a few minutes before 

 again being brought down to the water and reexamined. There is no more 

 common cause of the failure of the stains to take than the imperfect removal 

 of the wax. 



Only two solutions are required for staining. These are a 2.5 per cent solution 

 of ferric alum and a 0.5 per cent solution of hematoxylin. The only difficulty 

 in making the ferric alum solution is to secure a pure and unoxidized sam- 

 ple of the reagent. Most of the crystals in a new bottle are of a clear violet 

 color, but after a bottle has been opened for some time, particularly if the stop- 

 per is loose, most of the crystals become covered with a brownish deposit 

 which must be scraped off with a knife before the solution is prepared for 

 staining. If the brown powder on the outside of the crystal forms a layer of 

 any thickness, it is best to reject the whole and secure a fresh supply of the 

 reagent. Hematoxylin itself has little staining effect; the color is produced by 

 the formation of lakes with hematein, an oxidation product of the hematox- 

 ylin. It was customary in former times to prepare large quantities of solution, 

 which were kept with the stopper loose in the bottle for a period of at least 

 one month before use. For the purpose of Heidenhain's technique, however, 

 it is far more important that a small quantity of the ferric alum should be 

 carried over into the hematoxylin solution than that the latter should be aged. 

 The staining will be found both simpler and more effective if a few drops of 

 hematoxylin are placed in the iron alum solution and a few drops of the iron 

 alum solution are placed in the hematoxylin. Both solutions, of course, should 

 be filtered immediately before use if the finest slides are required because 

 chromosome figures in a rat can be obscured by a very small particle of dust. 



Next the slides are taken from distilled water and placed in the mordant so- 

 lution. It matters little how long they remain in this solution though the usual 

 directions call for keeping them there overnight. This varies with every type 



