Abstract: Soil enzyme plays an important role in matter cycle and energy flow of soil ecosystem. The analysis of soil enzyme activities is the basis of Soil Enzymology. The traditional soil enzyme methods can analysis soil enzyme activities, but cannot identify spatial and temporal patterns of soil enzyme activities. In-situ soil zymography is based on fluorescent substrates, and it can reflect the continuous spatial changes of soil enzyme activities at a microscale by in-situ two-dimensional visualization and distinguish hotspots and non-hotspots. In-situ zymography has advantages such as high accuracy, high spatial resolution and high temporal resolution. In this review, we summarized the operating principle, technical advantage and relevant research of in-situ zymography. Besides, we put forward the research possibilities of combining in-situ soil enzymology with the other technologies, which could promote the development and application of in-situ zymography in China and provide a theoretical reference for the future development of Soil Enzymology.