Gelatin Zymography: Detecting Gelatinase Activity as a Partner for Invadopodia-Mediated Invasion by Tumor Cells

Zymography is a technique for detecting, identifying, and quantifying enzyme activity through gel electrophoresis. Gelatin zymography is used to detect matrix metalloproteinase (MMP) activity (a class of zinc-dependent endopeptidases) such as MMP-2 and MMP-9. We describe a semiquantitative method, optimized for determining the activity of secreted gelatinases in cell culture conditioned medium via the digestion of gelatin embedded in a polyacrylamide gel. This assay can also be partnered with a quantitative "invadopodia degradation" assay, which detects the degradation of FITC-labelled gelatin by actin-rich membrane protrusions known as invadopodia. These projections are enriched in proteolytic enzymes, including MMP-2 and MMP-9 in glioblastoma cells, and are secreted to modify/degrade ECM proteins and facilitate tumor cell invasion.