The cell wall of Mycobacterium contains a large number of lipids surrounded by peptidoglycan, so Mycobacterium is generally not easy to color, and the traditional staining method has to be heated and prolonged staining time to promote its coloration. Once the mycolic acid in Mycobacterium combines with the dye, it is difficult to be decolorized by acidic decolorizing solution.

The acid-resistant staining solution (Auramine O-Rhodamine fluorescence method) belongs to the fluorescent staining solution, which does not need to be heated and is relatively safer than the acid-resistant heat staining solution. The staining principle is that after Auramine O-Rhodamine staining as well as re-staining at room temperature and examination by fluorescence microscope with ultraviolet source, the acid-resistant bacilli show bright yellow color while the other bacteria and the material in the background show dark yellow color, which can be examined with low magnification microscope, and therefore the acid-resistant bacteria can be identified more quickly. This method can be used for low magnification microscopy and therefore can identify the acid-resistant bacteria more quickly.