268 TECHNIQUE 



used again until the retention of the iodine color indicates that all 

 free mercury salt has been removed from the tissues. The tissue 

 then may be stored in 70 per cent alcohol. 



A modification of Zenker's fluid is called Hellys fluid. It has 

 the same formula as the above, but substitutes 5 cc. of formalin 

 for the acetic acid. It gives an excellent picture of protoplasmic 

 structure, but care must be taken in later steps in technique so that 

 distortion through shrinkage does not take place. Chromosomes 

 are not so distinct as with Zenker's fluid, but cytoplasmic structures 

 are well preserved and can be demonstrated by various stains. 

 Helly's fluid must be thoroughly washed out in running water 

 after twelve hours' fixation, and the tissues must also be later 

 immersed in 70 per cent alcohol to which iodine has been added. 



Formalin Solutions.— A 10 per cent solution serves as a good pre- 

 servative and fixative in the case of delicate tissues that will stand 

 hardening. It has been used as a preliminary reagent in a number 

 of techniques employed for nervous tissue. Tissues fixed in it 

 should be transferred to 70 per cent alcohol after twelve hours' 

 treatment with the formalin solution. Shrinkage often occurs later 

 during paraffin embedding. 



Carnoys Fluid.— The formula is: 75 cc. of 100 per cent alcohol 

 and 25 cc. of glacial acetic acid. This fixing fluid fixes chromosomes 

 well, precipitates glv'cogen, but usually dissolves chondriosomes and 

 the Golgi apparatus. It penetrates quickly, so that an hour serves 

 to fix soft pieces of 1 cm. in thickness. It combines the effects of 

 two chemicals, the alcohol causing precipitation of the proteins 

 and glycogen of the cytoplasm; the acetic acid fixing the nucleo- 

 proteins and preventing some of the shrinkage and hardening of 

 the alcohol. Tissues should be washed in absolute alcohol after 

 fixation before proceeding to embed. 



For additional information concerning fixatives the student is 

 referred to the texts cited at the end of this chapter, but for a 

 general treatment of fixation reactions it is recommended that 

 J. K. Baker's monograph on " ( \vtological Technique" be studied. 



Containers. As containers for the fixative and excised material, 

 it will be found advantageous to use wide-mouthed bottles of 1- or 

 2-()imce caj^acity i)rovided with cork stoppers. In order that the 

 fixative may reach all ])arts of the surface of the material, it is 

 well to place a small bit of absc^-bent cotton in the bottom of each 

 l>ottle to prevent the tissue from adhering to the glass, and so keep- 

 ing one surface from free contact with the fixing fluid. The bottle 



