SECTION TWO 



2. Fix sections to slides; dewax and bring down to distilled 

 water as usual. 



3. Stain with Ehrlich haematoxylin five to ten minutes; then 

 blue and wash in lithium carbonate saturated aqueous. 



4. Stain for fifteen to twenty-five minutes in the phloxine 

 solution; then wash with distilled water. 



5. Stain in aniline gentian violet for about ten minutes. 



6. Rinse in distilled water. 



7. Immerse in Gram's iodine solution for one minute. 



8. Wash in distilled water. 



9. Decolorize with aniline oil until the stain ceases to come out 

 of the sections. 



10. Rinse well in several changes of xylol and mount. 



Results: 



Branched forms are stained blue, while clubs appear red. 



Reference: MalIor>', F. B. (1938), p. 279. 



HAEMATOXYLIN, PHOSPHOTUNGSTIC 



For Pleuropneumonia organisms in sections of lung 



Solution required: 

 Phosphotungstic acid haematoxylin (Mallory). 



Technique: 



1. Pieces of tissue should be fixed in Bouin, Carnoy or absolute 

 alcohol and embedded in paraffin wax in the usual manner. 



2. Fix sections to slides ; dewax, pass through descending grades 

 of alcohol down to distilled water, as usual. 



3. Stain in Mallory's phosphotungstic acid haematoxylin in 

 a stoppered staining jar for twenty-four hours, without treatment 

 with the usual potassium permanganate and oxalic acid solutions. 



4. Pour off excess stain ; drain well without washing ; then care- 

 fully blot dry. 



5. Dehydrate quickly in absolute alcohol. 



6. Clear in xylol and mount. 



239 



