464 METHODS AND FORMULAS DS 23.12-DS 23.13 



23.12 Wolbach 1919 11343, 41:75 



REAGENTS REQUIRED! A. 70% alc. 98, ADS 12.2 Lugol (1905) 2; B. 0.5% sodium thio- 

 sulfate; C. distilled water 100, sodium bicarbonate 0.001, methanol 10, DS 13.13 

 Giemsa 1902 2; D. ADS 21.2 Wolbach 1919 2; E. acetone 70, xylene 30 

 method: [3 iU-5 m sections of Wolbach F 3700.0000 material, dewaxed and brought to 

 90% ale] -^ A, 20-30 mins. -^ 70% ale, wash — » water, wash -^ B, b mins. -^ wash 

 -^ C, 30 mins. — > C, fresh solution, 30 mins. — > C, fresh solution, overnight -^ D, 

 flooded on slide, 15-30 sees. — * E, till dehydrated -^ xylene — > cedar oil 

 result: nuclei and Rickettsiae, blue; cytoplasm, red. 



23.12 Zinsser and Bayne-Jones 1939 Zinsser and Bayne-Jones 1939, 654 



stock solutions: I. phosphate buffer pH 7.5 100, 40% formaldehyde 0.5; II. 1% 



methylene blue in methanol 

 reagents required: A. stock I 100, stock II 0.75, 40% formaldehyde 5; B. water 100. 



safranin 0.05, acetic acid 0.075 

 method: as DS 23.12 Casteneda 1930 



23.12 Zinnser, Fitzpatrick, and Hsi 1939 see DS 23.12 Macchiavello 1938 (note) 



23.13 METHODS FOR NEGRI, AND OTHER VIRUS INCLUSION, BODIES 



Negri bodies are generally supposed to be aggregations of virus particles, though they 

 may well be the breakdown products of the cell itself. The stains which have been de- 

 veloped for them are far more specific than the stains given for Rickettsiae in the 

 previous section. The method of Schleifstein 1937 is in general employment in the 

 United States today, although a far better demonstration, if speed is not of more 

 importance than accuracy, is to be obtained from the technitiue of Lepine 1935. The 

 "glyceric ether" mentioned in the technique of Manou^lian is not an article of com- 

 merce in the United States, but is so readily available in Europe, where Manoiielian's 

 method is widespread, that it is here included. Particular attention should be given to 

 the formula of Stovall and Black 1940, for it is the only one of the formulas which have 

 been given for the staining of these minute particles in which an acid buffer is emploj-ed. 



23.13 Barreto 1944 test. 1945 Conn 20540b, 20:66 



reagents required: A. either DS 11.121 Weigert 1903 or DS 11.123 Ehrlich 1886; B. 



1% hydrochloric acid in 70% ale; C. water 100, methyl blue 0.12, eosin 0.12; D. 95% 



alc. 100, picric acid 0.005, acetic acid 0.1 

 method: [sections] -^ water — > A, till stained — > B, till differentiated — > tap water, till 



blue — > C, on slide, 15 mins, -^ rinse — > D, till differentiated — » 90% alc. -^ balsam, 



via usual reagents 



23.13 Carpano 1916 test. 1942 Langeron Langeron 1942, 845 



reagents required: A. 1% eosin Y; B. DS 12.15 crystal violet 20, water 80; C. ADS 



12.2 Lugol (1905) 

 method: [5 fx sections of F 3700.0010 Zenker 1894 fixed material, or smears similarly 



fixed] —y A, 1 min. — > 95% alc, wash -^ B, warmed to steaming, 5 mins. — > C, used 



to wash B from slide, 1 min. — > 95% alc, till differentiated -^ balsam, via usual 



reagents 



23.13 Craigie 1933 11431,36:185 



reagents required: .4.2% mercurochrome; B. water 100, sodium phosphate, dibasic 

 (12 H2O) 0.35, mercurochrome 0.002, azur I 0.01, methylene blue 0.25 



PREPARATION OF B: Mix 1 2% mercurochrome, 57% Na2HP04.12H20, 1 1% azur I. Add 

 75 water and 25 1 % methj-lene blue. 



method: [air-dried blood smears or tissue scrapings] — > methanol, 5-10 mins. —* dry -^ 

 rinse — + blot —>■ A, 5-10 mins. —♦rinse — > blot —y B, 5-10 mins. — * quick rinse -^ dry 



recommended for: Paschen bodies. 



23.13 Coles 1935 11360,55:249 



reagents required: .4. DS 13.13 Giemsa 1902 (dry .stock) 0.75, glycerol 25, 95% alc. 



75; B. water 100, tannic acid 5, orange G I 

 method: [smears] — > A, 24 hrs. -^ wash -^ dry — * B, few moments — * wash -^ dry 

 recommended for: virus inclusions in general. 



