In Situ Zymography and Localization of Bright Green Fluorescent Gelatinase Activity in Tissue Sections

Abstract

In situ zymography is an efficient and low-cost technique to detect the activity of the gelatinases matrix metalloproteinase (MMP)-2 and MMP-9 in various tissues from humans and experimental animal models. This technique also shows the precise localization of gelatinases in large or small tissues and in cells. It is a one-day technique that uses a dye-quenched fluorescent gelatin (Dq-gelatin). The Dq-gelatin is loaded into tissue sections and upon its interaction with local active gelatinases, it is proteolyzed into highly fluorescent peptides that emit bright green fluorescence, thus reflecting the level of activity and localization of the gelatinases. Combined with other biochemical techniques in tissue homogenates and fractions, the in situ zymography provides a fluorescence microscopy and digital imaging technique to assess the subcellular localization and the role of gelatinases in tissue remodeling in health and disease.

Keywords: Extracellular matrix; Gelatin; Matrix metalloproteinases; Remodeling; Vascular.