In acid-fast staining, what is the role of the waxy cell wall in some bacteria?

The waxy cell wall in some bacteria, particularly those from the Mycobacterium genus, plays a crucial role in acid-fast staining by retaining the primary stain despite the application of decolorizers. This characteristic is primarily due to the presence of mycolic acids, long-chain fatty acids that create a hydrophobic environment. When the primary stain, typically carbol fuchsin, is applied, it penetrates the waxy cell wall. Following the decolorization step, which generally involves the use of an acid-alcohol solution, these bacteria still retain the primary stain because the waxy cell wall prevents the stain from being washed out. This property is what makes acid-fast bacteria appear red under microscopy, distinguishing them from non-acid-fast organisms, which lose the stain and take up the counterstain, appearing blue. This critical differentiating feature is utilized in clinical microbiology to diagnose infections caused by these resistant bacteria, such as tuberculosis.