254 Cytoplasmic Elements (chap. 18) 



2. Desaturate the solution by: {a) dilution of a saturated solution with equal 

 volumes of 60% alcohol, or {b) by refrigeration. When the solution has 

 been refrigerated long enough to have acquired the refrigerator tempera- 

 ture, filter. Either method does not alter the staining quality of the solu- 

 tion and produces negligible amounts of precipitate. 



Oil Red O for Fat 



fixation: 10% formalin or other aqueous general fixatives. 



solutions: 



Oil red O stock solution: 



oil red O, C.I. 26125, saturated in 99% alcohol (250-500 mg. 

 dye/ 100 ml. alcohol) 



Working solution: 



stock solution 6.0 ml. 



distilled water 4.0 ml. 



Let stand 5-10 minutes (no longer). Filter. Use immediately. 



Hematoxylin, see page 125. 



procedure: 



1. Frozen sections, 15 microns. 



2. Place in distilled water. 



3. If HgClo is present, treat with Lugol's and thiosulfate; wash. 



4. Stain in oil red O: 10 minutes, in closed container to reduce evap- 

 oration. If dye begins to settle out before 10 minutes have elapsed, 

 remove sections to water, or precipitate inay settle on sections. 



5. Wash in water. Make transfers with glass rod and wipe it clean 

 after each section. 



6. Stain in Mayer's hematoxylin (or like): 2-3 minutes. 



7. Wash in tap water: 3 minutes. 



8. Blue in Scott's solution (page 412): 3 minutes. 



9. Wash in tap water: 5 minutes. Mount in gum syrup or glycerol 

 jelly. For permanency, ring cover glass. 



results: 



fat — orange-red or brilliant red 

 nuclei — blue 



Oil Red O with Tween 40 (bell, 1959) 



fixation: Baker's formalin recommended (page 14), or other general 

 aqueous fixative, no alcohol. 



