The assay is designed to measure either MDA alone (in hydrochloric acid) or MDA in combination with 4-hydroxyalkenals (in methanesulfonic acid). It is based on the reaction of a chromogenic reagent, N-methyl-2-phenylindole with MDA and 4-hydroxyalkenals to yield a stable chromophore with maximal absorbance at 586 nm.

Lipid peroxidation is a well-established mechanism of cellular injury in both plants and animals, and is used as an indicator of oxidative stress in cells and tissues. Lipid peroxides are unstable and decompose to form a complex series of compounds including reactive aldehydes.

Polyunsaturated fatty acid peroxides generate malondialdehyde (MDA) and 4-hydroxyalkenals (HAE) upon decomposition. Measurement of malondialdehyde and 4-hydroxyalkenals has been used as an indicator of lipid peroxidation.