478 METHODS AND FORMULAS DS 23.213-DS 23.214 



23.213 Pottenger 1942 test. 1942 Farber 20540b, 17:183 



REAGENTS REQUIRED: A. DS 11.43 Pottenger 1942; B. 3% hydrochloric acid; C. 0.3% 



picric acid 

 method: [heat-dried, fixed smear] — > A, 15 mins. 65°C. — > B, 30 sees. — > 70% ale, till 



no more color comes away -^ C, 10 sees. — > rinse — > dry 



23.213 Randolf and Mikele 1944 665, 49:109 



STOCK solutions: I. 1% magenta in propylene glycol. II. 5% phenol 



REAGENTS REQUIRED: A. stock I 20, stock II 80; B. 1% hydrochloric acid in 70% ale. 



C. 1% methylene blue 

 method: [heat-fixed smears] -^ A, 4 mins. —> wash — > B, till decolorized — » C, 1 min. — ' 



wash -^ dry 



23.213 Spehl 1918 see DS 23.213 (note) Spengler 1907 



23.213 Schulte-Tigges 1920 7276,46:1225 



REAGENTS REQUIRED: A. DS 11.43 Ziehl 1882; B. 10% sodium sulfite; C. sat. aq. sol. picric 



acid 

 method: [heat-fixed smears]—* A, warmed to steaming, 1 min. — > wash -^ B, till de- 

 colorized —f wash -^ C, few mins. — > wash -* dry 



23.213 Spengler 1907 7276, 33 :337 



REAGENTS REQUIRED: A. DS 11.43 Ziehl 1882; B. 95% ale. 50, sat. sol. {circ. 1.2%) picric 



acid 50; C. 15% nitric acid 

 method: [heat-fixed smears] -^ A, warmed to steaming, 1 min. — > B, mixed with A on 



slide, few sees. -^ 70% ale, wash -* C, 30 sees. — > 70% thorough wash -^ B, 15 sees. 



-^95% ale, quick rinse — > dry 

 note: Spehl 1918 (6630, 81:248) substitutes a mixture of 3 parts DS 11.43 Ziehl 1882 



with 2 parts DS 11.44 NicoUe 1871 for A above. 



23.213 Weiss 1942 665, 46:199 



REAGENTS REQUIRED: A. ADS 12.1 Weiss 1942; B. 2.5% safranin; C. 15% acetic acid in 



acetone; D. DS 11.44 Loffler 1890 

 method: [flame-fixed smears]—* A, on slide, heated to steaming, 5 mins. -^ wash -^ 



B, on slide, heated to steaming, 5 mins. — * wash -^ C, till decolorized -^ wash -^ D, 

 1 min. —> wash — * dry 



23.213 Ziehl 1882 see DS 23.213 Neelsen 1883 



33£14 Methods for Spirochetes 



Spirochetes are resistant to dye-staining techniques, and are usually better demon- 

 strated either unstained with dark ground illumination, or else with the aid of one of 

 the metal-staining techniques given in Chapter 23. When they are to be stained with 

 dyes, most of the early techniques involve the utilization of Giemsa's 1902 technique 

 (Chapter 20 DS 13.13), usually with the addition of a strongly alkaline differentiating 

 solution, with or without prior mordanting or subsequent differentiation in a solution 

 of tannic acid. More recently, certain techniques have appeared such as that of Noguchi 

 1921, in which prior mordanting is conducted in a solution buffered to pH of 7.6, with 

 subsequent staining in any stain desirable to the technician. A very unusual and ex- 

 cellent stain, which should possibly have been placed in the section on metal staining, 

 is that of Ono 1938. He utilized the heavy deposition of oxides of manganese from a 

 solution of potassium permanganate, after mordanting with formaldeh_yde. This method 

 tends to cause darkening of the background, because the oxides of manganese will be de- 

 posited on an)^ organic detritus present, but if the culture is relatively free of organisms, 

 or materials other than the spirochetes, the method can be confidently recommended. 



23.214 Becker 1920 test. 1928 Schmorl Schmorl 1928, 400 



REAGENTS REQUIRED: A. F 0000.1010 Ruge (1942); B. water 100, tannin 10, phenol 1; 



C. DS 11.43 Ziehl 1890 



method: [smears] — > 1 min. — » rinse -^ B, 30 sees. 40°-50°C. — > rinse -* C, warmed to 

 steaming, 30-45 sees. — > wash — > dry —> balsam 



