110 Biological Stains 



J9 AZURE B 



Azure B is the tri-methyl derivative of thionin : 



CH3 CH3 



\ / 



N_/\_S _/\_N-H 



/ I I I I \ 



CH3 III CI 



\/~N=\/ 



C15H16N3SCI; Mol. Wt. 305.819 

 {A basic dye; absorption maximum 648-653) 



Like azure A, this compound was first recognized by Kehrmann 

 (1906) as a constituent of polychrome methylene blue. It can 

 easily be prepared in impure form by oxidation of methylene blue, 

 and is now available if anyone desires to obtain it. 



Curve 3 in Fig. 12, p. 108, shows its spectral characteristics. 

 It is distinctly less red than any of the other azures. 



MacNeal (1925) stated that its staining effects were much the 

 same as could be obtained with a mixture of azure A and methylene 

 blue, and that it therefore could be of little importance in poly- 

 chrome methylene blue. His statement has generally been inter- 

 preted as meaning that azure B is not a satisfactory stain. Holmes 

 and French (1926) also stated that this dye has little staining 

 value; but their conclusions were based upon work with an impure 

 sample, the performance of which was so poor as not apparently to 

 justify further purification. Haynes' later work (1927) on sections 

 of fixed tissue, with this same sample of azure B, agreed par- 

 tially with this, although she found the dye, if used in buffered 

 solution, rather better than fairly pure methylene blue in fresh 

 solution. Jordan and Heather (1929) definitely specify this dye in 

 a stain for Negri bodies. Moreover, recent tests by the Stain 

 Commission of certain exceptionally good Giemsa stains indicate 

 the possible presence of more azure B than in less satisfactory 

 batches. Roe, Lillie, and Wilcox (1940), in fact, have concluded 

 that azure B is one of the most important constituents of Giemsa 

 stain, especially when used for staining malaria parasites in thick 

 blood films. This work has been followed up by a series of other 

 papers by Lillie and his associates, and they have distributed 

 samples to other laboratories for testing; as a result it seems diffi- 

 cult to question their conclusions. Accordingly it is now recog- 

 nized that Azure B is one of the most important constituents of 

 ''Azure I". 



