350 Histochemistry (chap. 21) 



(perform Prussian blue reaction) or calcium deposits (remove before 

 incubation in citrate buffer of pH 4.5 ±: 10-20 minutes). 



George and Thomas {I960) coat sections with thin layer of 5-10% 

 aqueous solution of pure gelatine (preserve with thymol not phenol). 

 Place in refrigerator freeze chamber to solidify coating. Place in 6% 

 cold neutral formalin to harden. Wash and proceed to incubate. 



Eapen {I960) proved that lipase is activated by gelatine and coated 

 sections to give better results. 



Succinic Dehydrogenase 



The succinic oxidase system is responsible for the oxidation of the 

 majority of tissue substrates and is a part of the oxidation-reduction 

 system in tissues. It is widely distributed, particularly in heart muscle 

 and kidney. 



Succinic dehydrogenase is demonstrated by the reduction of tetra- 

 zolium salts. The tetrazolium (colorless) acts as a hydrogen acceptor and 

 becomes reduced to a blue water-insoluble pigment, diformazon. 



Tetrazolium salts: 



TFT— 2,3,5-triphenyl tetrazolium chloride: simplest, with limited sen- 

 sitivity. 



BT — 2,2',5,5'-tetraphenyl-3,3' (3,3'-dimethoxy-4,4'-biphenylene) ditetra- 

 zolium chloride: better pigment qualities. 



Nitro BT— 2,2Mi-p-nitrophenyl-5,5'-diphenyl-3,3'-(3,3'-dimethoxy-4,4'- 

 biphenylene) ditetrazolium chloride. 



Most methods require anerobic (absence of oxygen) conditions to pre- 

 vent atmospheric oxygen from competing with the tetrazolium for the 

 electrons of the dye and thus decrease reduction. Sodium succinate is 

 the substrate. 



Sections must be fresh frozen, cut at 30 microns (not less than 20 mi- 

 crons) and are improved by cutting in a cryostat. They may be floated 

 on a beaker of the incubating fluid inside the cryostat or moinited on 

 slides and air dried in the cryostat. For anerobic conditions remove air 

 from the solution with vacuiun, or boil the substrate and then cool to 

 37 °C. In either case, keep in fidl, tightly closed bottles (small weighing- 

 bottle are excellent). Sections are dropped in and the bottle immedi- 



