Fill Staining 257 



allow the water to dissolve the crystals. This method eliminates pos- 

 sibility of breathing the fumes. 



procedure: 



1. Frozen sections, 15 microns, into water. 



2. Transfer to osmic acid solution: 24 hours. 



3. Wash in several changes distilled water: 6-12 hours. 



4. Treat in absolute alcohol: 4-5 hours, for secondary staining of fat. 



5. Wash well in distilled water: 5 minutes or more. 



6. Moinit in glycerol jelly. 



results: 

 fat — black 

 background — yellowish brown 



Naphthalene Yellow (sills and marsh, 1959) 



Naphthalene yellow in 60% acetic acid was used to restore the yellow 

 color to the fat of gToss formalin-fixed specimens. This cannot be used 

 for sections; the color is too pale. 



Fluorescent iVIethod (\[etcalf and patton, 1944; peltier, 1954) - 



fixation: 10% formalin (salts of heavy metals — ^HgCU — have a quench- 

 ing effect on fluorescence), or use fresh tissue. 



solution : 



Phosphine 3R: 



phosphine 3R^ 0.1-1.0 gm. 



distilled water 100.0 ml. 



procedure: 



1. Cut frozen sections, 10 microns. 



2. Wash in distilled water. 



3. Stain in phosphine solution: 5 minutes. 



4. Rinse quickly in distilled water. 



5. Moiuit in glycerol, or examine as a water mount. 



results: 



lipids (except fatty acids, cholesterol and soaps) will fluoresce brilliant 

 white 



comment: 



For details concerning fluorescent microscope and equipment, see 

 page 102. 



* George T. Gurr, London, England; or Pfatz and Bauer, Inc., Empire State Bldg., XA'. 



