Phospholipases A2 (PLA2) are hydrolytic proteins, which cleave fatty acid off the second position (sn-2) of the phospholipid. An increased activity of PLA2 correlates with the course of many different inflammatory processes in the body. For the purpose of diagnosing and predicting pathological processes, systems for detecting the PLA2 activity are being developed. The key component of all test systems is a substrate of lipid or non-lipid nature, the breakdown of which by the enzyme leads to the appearance of analytical signal. Lipids as such do not absorb light in the visible region and do not fluoresce. Therefore, to determine the activity of PLA2, substrates with various labels are developed. Test systems for determination of the PLA2 activity can be divided into three groups, depending on the stage of the enzyme action a signal is formed at: (1) systems based on the detection of hydrolysis products; (2) systems based on the cleavage of fluorescently labeled substrates, and (3) systems based on the detection of membrane disintegration. Each of these groups has its own requirements for the structure of the substrate. This review is focused on the structure of PLA2 substrates used in systems to determine the enzyme activity; the proposed classification allows one to identify the strengths and weaknesses of existing detection systems and will be relevant when designing new test systems.