280 APPENDIX. 



2. Dehydration. After washing for twelve to twenty-four hours in running 

 water, following corrosive sublimate fixation, or simply "washing for a few minutes 

 after formalin, the tissues should be placed in 70% alcohol. They may be kept- in 

 this indefinitely. If they are to be sent to a laboratory for sectioning, it is advisable 

 lo moisten a pledget of cotton in 70% alcohol and fill in the bottom of the bottle with 

 it. Then drop in the tissues and pack in gently over them sufficient 70% alcohol 

 saturated cotton to fill up the bottle. All the alcohol should be absorbed by the 

 cotton so that if the bottle should break in transit there would be no damage from 

 the alcohol. The stopper of the bottle should be paraffined or sealed with wax. 



Tissues may be left in the 70% alcohol 1 2 to 24 hours and should then be trans- 

 ferred to 95% alcohol for an equal time. They are then transferred to absolute 

 alcohol, where they remain from 2 to 12 hours and are then placed in xylol. The 

 time in xylol should be as short as possible. So soon as the tissue looks clear it 

 should be removed 30 minutes to two hours. 



3. Imbedding. The tissue is now transferred to melted paraffin. Paraffin 

 melting at 48 C. for winter work, and that melting at 54 C. for summer. 

 The time in the paraffin should not he prolonged. Two hours will ordinarily suffice. 

 Some leave in the paraffin for 12 to 24 hours. 



Next take a paper box (made of stiff writing-paper folded over a square of wood) 

 and fill with the melted paraffin. As quickly as possible drop in the piece of tissue 

 taken out of the paraffin bath with heated forceps and, so soon as the paraffin begins 

 to solidify on the surface, place the paper box in ice water. When paraffin is rapidly 

 cooled, crystallization is less. 



The Acetone Method. Take the tissues out of the 70% alcohol and place in 

 acetone. After remaining in acetone for one to two hours, the tissues should be 

 transferred to fresh acetone for an equal length of time. They should then be 

 placed in xylol for about one-half hour and then imbedded in paraffin as directed 

 above. 



The Chloroform Method. The procedure may be the same as in the method of 

 passing through alcohols to xylol, substituting chloroform for xylol and then trans- 

 ferring to paraffin. 



Where absolute alcohol is not obtainable, very satisfactory results may be obtained 

 by transferring to a mixture of 95% alcohol and chloroform after immersion in 95% 

 alcohol. Then going from the alcohol-chloroform mixture to pure chloroform 

 thence to paraffin. 



When a piece of tissue is not more than one-fourth inch square and one-eighth 

 inch thick, it is very easy to run it through in three to six hours. Thus: 



10% Formalin (in 37 C. incubator), i hour. 



70% Alcohol (in 37 C. incubator), i hour. 



95% Alcohol (in 37 C. incubator), i hour. 



Absolute Alcohol (in 37 C. incubator), 1/2 hour. 



Xylol (in 37 C. incubator), 1/2 hour. 



Paraffin (in 55 C. incubator), 1/2 to 2 hours. 



