In situ zymography

Zymography A method for detecting enzyme activity on a matrix, usually a polyacrylamide gel or agarose gel after electrophoretic separation. See Frederiks, W.M. and Mook, O.R., Metabolic mapping of proteinase activity with emphasis on in situ zymography of gelatinases review and protocols, J. Histochem. Cytochem. 52, 711-722, 2004 Lombard, C., Saulnier, J., and Wallach, J., Assays of matrix metalloproteinases (MMPs) activities a review, Biochemie 87, 265-272, 2005. 

In situ zymography kit (EnzChek Gelatinase Assay Kit, Molecular Probes, Eugene, OR). 

Examples of the results obtained after EB injection (Figs. 3c and 6), in situ zymography (Fig. 4b), or direct IMF visualization of extravasated IgGs (Fig. 7) are shown below. 

Fig. 4 In situ zymography and immunohistochemistry of MMPs Relationship with BBB disruption induced by HIV-1 gpl 20. (a) Injection of gpl 20 into the rat CP induced upregulation of matrix metalloproteinases (MMPs). Frozen cryostat sections of CP injected with 500 ng gpl 20 and stained by in situ zymography. Left column The fluorescent product was observed in blood vessel-like structures and in cells. Middle column-. Positive control. In situ zymography detects intense fluorescence in the hippocampus. Right coiumrr. No gelatinolytic activity was seen In the contralateral unInjected side or in CPs injected with saline (not shown), neither when a section of the test brain was incubated with 1,10-phenanthroline, a nonspecific inhibitor of MMP activity, (b) There was a colocalization between In situ zymography and RECA-1. (c) Colocalization between MMP-2 and the endothelial cell marker RECA-1 In CPs Injected with gpl 20. MMP abnormalities were associated with EB leakage (not shown in this panel) (modified from [25,48,105])...

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