Chapter VI: The Paraffin Method 53 



the microscope. When in a dividing cell the chromosomes become 

 sharply denned, the decolorization should be stopped. 



8. Wash in several changes of water for 2 to 3 hours. If any 

 of the iron-alum is left in the sections the color will fade later. 



9. Wipe off the excess of water, transfer the slide to 95 per 

 cent, alcohol, followed by absolute alcohol and carbol-xylol. 



10. Mount in balsam. 



NOTE. Iron hematoxylin is perhaps the one most important stain in 

 use today. The student should practice the method until he has 

 mastered it. 



It is better though not absolutely essential that the stain be " ripe." 

 If the stain is to be simply for general histological instead of cytological 

 work, the baths may be curtailed considerably. For example, immersion 

 for 30 minutes in the iron solution, then for 45 minutes in the stain fol- 

 lowed by very brief differentiation in the iron solution, yields a good 

 general preparation but the finer details of cell structure (centrosomc, 

 etc.) are not brought out. 



V. BORDEAUX RED AND II^ON-ALUM HEMATOXYLIN 



j 



Use the sections which were reserved for this method. The 

 method is identical with the one just outlined, except that between 

 step 2 and step 3 the following directions should be inserted: 2a, 

 transfer the sections from water to Bordeaux red for 2 hours 

 (12 hours will do no harm), then wash them in water and proceed 

 to step 3. 



NOTE. Before proceeding further, kill a female cat or rabbit to 

 secure tissues for the celloidin method and to correct failures in the 

 paraffin method. In addition to the tissues specified before, prepare 

 (fix in Gilson) pieces of tendon, cartilage, spleen, lymph gland, pancreas, 

 and salivary glands. (If the reagents are at hand and time permits, 

 the student, indeed, might advantageously prepare a number of tissues 

 according to the methods indicated in Appendix C.) Fix the ovary in 

 Gilson, and reserve it for the paraffin method for delicate objects. Fix 

 parts of the brain and cord in Erlicki and in formalin as previously 

 indicated, and place bits of muscle in which nerves terminate plentifully 

 (e.g., intercostals) in formalin. Larger pieces (up to 2 cm.) may be 

 used of such tissues as are to be imbedded in celloidin. Bear in mind 

 that the larger the tissue the longer must it be left in the different 

 reagents. Select the necessary parts of the digestive tract to prepare 

 longitudinal sections in celloidin from esophagus to stomach and from 

 stomach to intestine. As soon as possible begin the preliminary steps 

 in the celloidin method (chap, vii) so that there may be no loss of time. 



