Auramine-O Staining
Auramine is preferred to Ziehl-Neelson staining due to its sensitivity, and is rapidly becoming the standard method of diagnosing M. tuberculosis and other infections in most laboratories.
It is a very simple protocol, although it does require a fluorescent microscope and slightly more expensive reagents.
Procedure
Smears are first airdried and fixed, using either methanol (3-5 min) or heat.
Auramine-O (1:1000 solution, in ethanol, phenol, and distilled water) is flooded onto the slides to stain mycolic acid.
After 15 minutes, the slides are washed with tap water and then decolorized using HCl (0.5% in ethanol).
After 3-5 minutes, slides are again washed in tap water before being counterstained with potassium permanganate (0.5%) for 30-60 seconds.
Slides are washed for a last time and airdried for 10-45 minutes.
Slides are examined using a fluorescent bulb, using the following scale: