80 CYTOCHEMISTRY OF ALDEHYDES 



Standard Technique 



In any detailed investigation it is, of course, necessary to em- 

 ploy a considerable number of variations in technique. But it is 

 convenient in making preliminary investigations to have a stand- 

 ard technique. The technique which will be given demonstrates 

 the sum of free aldehyde and acetal aldehyde. Frozen sections 

 must be used. The normal procedure of infiltration with wax 

 will remove practically all the free fatty aldehyde and fatty 

 acetal aldehyde. It is this which enables the Feulgen technique 

 for thymonucleic acid to be carried out without masking by other 

 aldehydes. The steps for demonstration of the sum of acetal 

 and free aldehyde are as follows: 



1. Sections of tissue not more than 2 millimeters in thickness are fixed 

 in a solution containing 8 percent formaldehyde and 5 percent acetic acid. 



2. After a minimum of 2 hours' fixation, frozen sections are cut. 



3. The sections are washed with distilled water and then placed in cold 

 0.1 N hydrochloric acid for not more than 15 minutes. This liberates 

 aldehyde from any acetal aldehyde present. 



4. The sections are washed in distilled water to remove hydrochloric 



acid. 



5. Sections are transferred to reduced fuchsin for 15 minutes. 



6. The sections are given three washings, each of 5 minutes, in a solu- 

 tion containing sulfur dioxide. The sulfur dioxide solution can be pre- 

 pared conveniently by mixing 10 milliliters of 10 percent sodium bisulfite 

 solution with 10 milliliters of N hydrochloric acid, and adding distilled 

 water to 200 milliliters. 



7. The sections are washed in distilled water and mounted in glycerol 

 or Canada balsam. 



The fixative has been chosen because it gives a reasonable 

 compromise between good fixation of cytoplasm and good fixa- 

 tion of the nucleus in many tissues. Other tissues may need a 

 modified form of this fixative: for example, it may perhaps be 

 necessary to make the fixative up in sea water for the tissues of 

 marine animals. The high formaldehyde content is valuable in 

 protecting free aldehyde against oxidation. The whole procedure 

 up to step 5 can also be carried out when necessary under an- 

 aerobic conditions. This is most conveniently done by carrying 

 out the treatment with fixative, etc., with the material in a 

 Thunberg tube. The frozen sections can, if necessary, be cut 

 in a closed box in an atmosphere of nitrogen. It is best to cut 

 sections as soon as possible after fixation is complete. With 



