MEDICAL AND BIOLOGICAL STAINING TECHNIQUES 



ALCIAN BLUE 

 For bacterial polysaccharides and capsules 



Note: This dye will stain bacterial capsules and insoluble polysac- 

 charides in both pure and mixed cultures of bacteria and protozoa. It 

 is specific for bacterial polysaccharides, and enzymatic degradation 

 of these results in the loss of this stain. 



Solutions required: 



A. Alcian blue . . . . . . . . i gm. 



Alcohol 95% ,. .. .. 100 ml. 



B. Solution A . . . . . . . . i volume 



Distilled water . . . . • • 9 volumes 



Note: This solution which deteriorates 

 within a few days, should only be prepared 

 as and when required for immediate use. 



C. Carbol fuchsin (Ziehl-Neelsen) 



Technique: 



1 . Fix air-dried smears by flaming in the usual way. 



2. Stain with solution B for one minute. 



3. Pour off excess stain and wash with water; then allow the 

 preparation to dry in the air. 



4. Counterstain with carbol fuchsin for a few seconds. 



5. Wash immediately with distilled water to prevent over- 

 staining. 



6. Allow the preparation to dry in the air ; then examine. 



Results: 



Capsule and other bacterial polysaccharides are stained blue: 

 cellular material, red. 



Note: Various carbohydrates including adonitol, arabinose, cello- 

 biose, dextrin, glucose, inositol, inulin, mannitol, mannose, rhamnose, 

 sorbitol, trehalose, xylose, and certain enzymes including papain, 

 pepsin, rennin gave a positive reaction with alcian blue. 



The fact that the internal polysaccharides of the cell remain 

 unstained is attributed to the complexity of the dye molecule 

 which prevents its penetration of the cell wall. 



Reference: McKinney, Ross E. (1953), J^. Bact., (U.S.A.), 66, 453-4, ** Stain- 

 ing bacterial polysaccharides. " 



