The PHB staining is used to detect the presence of the prokaryotic storage compound poly-3-hydroxybutyrate. This storage material accumulates within bacteria under stress conditions and is considered a value added product as it can be used in many industrial appplications.
Solution (1): Sudan Black B (IV) 0.3% w/v in 60% ethanol
Solution (2): Safranin 0 0.5% w/v aqueous
1. Add a diluted sample onto the microscopic slides and allow to air dry. (Do not heat fix)
2. Add Solution 1 onto slide for 10 min and briefly rinse with water for 10 seconds.
3. Stain with Solution 2 for 10 seconds, rinse well with water and blot dry.
Upon viewing at 1000x magnification, the presence of black intracellular granules is a positive indication of stored PHB.