Oil Red O stain is based on the use of a lysochrome (lipo-soluble dye) and an azo dye used for the visualization of neutral triglycerides and lipids on tissue sections.

Oil Red O is one of the Sudan dyes such as Sudan III, Sudan IV and Sudan Black B. However, Oil Red O has gradually replaced the Sudan III and Sudan IV stains because Red coloration produced by the Oil Red O is much more intense, which facilitates visualization.

Oil Red O stain is done on fresh or frozen samples. The tissues included in paraffin are not compatible with this coloration because the preparation of cuts and especially the use of solvents eliminate lipids.

Oil Red O stain is based on the use of 4 reagents for the staining of fat cells according to Johnson's method.