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Fig. 5 Differential in vivo zymography with two peptides displaying differential susceptibility to MMP-mediated hydrolysis. This embryo was injected with a mixture of two MMP substrates (substrate I and substrate VI), whose breakdown products fluoresce in either the red or green channel, respectively. Both substrates are susceptible to hydrolysis by MMPs 2, 9, 13, and 14, but with different relative and absolute efficiencies. In the red channel, the fluorescent breakdown products of MMP-substrate peptide I are observed primarily in the brain, the vitelline circulation, squamous epithelial cells scattered across the surface of the embryo, the hatching gland, cells in the dorsal neural tube, maturing myotome boundaries, and the perichordal sheath. In the green channel, the fluorescent breakdown products of MMP-substrate peptide VI are observed primarily in the hatching gland, brain, myotome boundaries, cells of the dorsal neural tube, and the perichordal sheath. This suggests that sites where both peptides are similarly degraded (brain, myotome boundaries, cells of the dorsal neural tube, and notochord sheath) may harbor active MMPs 2 and/or 9, to which both peptides are similarly susceptible. Sites where one substrate is degraded significantly faster than the other must harbor protease(s) that preferentially degrade one of these substrates. In the scattered squamous epithelial cells and the cells in the vitelline circulation where substrate I is preferentially degraded, it is likely that MMP14, to which substrate I is more susceptible, is active. In contrast, in the hatching gland, where substrate VI is preferentially degraded, it is likely that MMP13, to which it is more susceptible, is the dominant protease. (Color figure is available at liebertonline.com.)