474 METHODS AND FORMULAS DS 23.211-DS 23.212 



23.211 Manson 1890 for formula see DS 11.44 Manson (1890); for method see DS 23 211, 

 Ziehl 1882 



23.211 Pick and Jacobson test. 1942 Langeron cii. Morax 



Langeron 1942, 1198 

 formula: water 100, DS 11.43 Ziehl 1882 3, sat. ale. sol. (circ. 2%) methylene blue 1.5 

 method: [heat-fixed smear] -^ stain, 20-30 sees. -^ wash -^ dry 



23.211 Sahli 1885 for formula see DS 11.44 Sahh 1885; for method see DS 23.211 Ziehl 

 1882 



23.211 Spehl 1927 6630, 107 :920 



REAGENTS REQUIRED: A. DS 12.15 gentian violet; B. sat. aq. sol. iodine; C. sat. aq. sol. 



picric acid; D. chloroform; E. DS 12.15 magenta 10, water 90 

 method: [dry smears] -^ A,2 min. -^ quick rinse -* B, 30 sees. -^ C, 30 sees. -^ dry -^ 



D, till no more color comes away -^ dry -^ E, 1 min. — » wash -^ dry -^ balsam 



23.211 Unna 1892 for formula see DS 11.44 Unna 1892; for method see DS 23.211 Ziehl 



23.211 Volkonsky 1933 for formula see DS 11.44 Volkonsky 1933; for method see DS 23.211 

 Ziehl 



23.211 Ziehll882 7276,8:451 

 REAGENTS REQUIRED: A. DS 11.43 Ziehl 1882, sol. A. 



method: [heat, or ale., fixed smears] -^ A, poured on slide, 30 sees. -^ water, wash — > dry 

 note: The same technique may be employed with DS 11.44 Loffler 1890, Manson (1929), 



Martinotti 1910, Sahli 1885, Unna 1892, Volkonsky 1933. See also DS 22.213 Neelsen 



1883. 



SS.212 Iodine Differential Methods 



The stains given in this section are commonly referred to as "Gram stains," and the organ- 

 isms which retain the color after any of the techniques here given are known as Gram-posi- 

 tive. Certain methods, such as Burke 1922, involve after-staining, usually with safranin, in 

 such a manner as to render those bacteria from which the stain is removed (the Gram-nega- 

 tive bacteria) red, or such color as will contrast well with the Gram-positive bacteria. The 

 methods require, without exception, preliminary heavy overstaining of the smear, the re- 

 moval of the stain with iodine, and usually, the removal of the iodine with acetone before the 

 application of the counterstain. 



23.212 Atkins 1920 11056, 5:321 

 reagents required: A. DS 11.45 Atkins 1920; B. ADS 12.2 Atkins 1920 



method: [heat-fixed smear] -^ .4, 1 min. —> wash —> J5, 1 min. — > wash — » 95 % ale, 

 rinse — > dry 



23.212 Burke 1922 11056, 7:159 



REAGENTS REQUIRED: A. 1% Crystal violet; B. 5% sodium bicarbonate; C. ADS 12.2 



Burke 1922; D. ether 25, acetone 75; E. 2% safranin O 

 method: [smears] — ♦ A, flooded on slide — > add 2-3 drops 5 to A on slide, 2-3 mins. — >• 



C, rinse -^ C, fresh solution, 2 mins. — > water, wash -^ blot —* D, dropped on slide til! 



no more color comes away -^ dry — * E, 10 sees. -^ water, wash -^ dry 

 result: Gram-positive, blue; Gram-negative, red 

 note: Kopeloff and Cohen 1928 (20540b, 3:64) substitute equal parts acetone-ale. for D 



above. 



23.212 Hucker and Conn 1923 20936, 93:1 



REAGENTS REQUIRED: A . BS 23.211 Hucker (1929); B. ADS 12.2 Gram 1884; (7. 95% ale. 



10, safranin O 0.25, water 100 

 method: [smears] —> A, 1 min. — ^ water, wash -^ B, 1 min. — » water, wash — * blot — ♦ 



95% ale. 30 sees. -^ blot — > C, 10 sees. — > water, wash — > dry 

 result: Gram-positive bacteria, blue; Gram -negative, red. 



23.212 Gram 1884 8645,2:185 



REAGENTS REQUIRED: A. DS 12.15 Crystal violet; B. ADS 12.2 Gram 1884; C. absolute 

 ale. 



