Novex™ 10% Zymogram Plus (Gelatin) Protein Gels, 1.0 mm, 10-well, 10 gels (1 box) - FAQs

Some small proteases work better if the sample is heated for a short time before loading so the protease is denatured better and it doesn't digest the casein on its way through the gel. This may also help to improve the band sharpness.

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Zymogram gels can be stained using Colloidal Blue stain or SimplyBlue SafeStain. The fixing step is not required. We recommend staining with SimplyBlue SafeStain after renaturing and developing the gel for enzyme activity. If greater contrast is needed for detecting weakly active enzymes, a 0.5% Coomassie R250 stain may be used.

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Zymogram gels are essentially Tris-Glycine gels containing the substrate. Protein standards run based solely on the percentage of acrylamide and hence should run the same in both kinds of gels. It is quite possible though that if the standard is pre-stained, the proteins will appear a different color because of the staining (or pre-staining) of the Zymogram gels.

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No, samples should not be heated or reduced so that multiunit proteases migrate as a single unit that can be renatured after electrophoresis. The sample should be one part sample and one part 2X Tris-Glycine SDS Sample buffer. Let the sample stand at room temperature for 10 minutes and then load the sample onto the gel.

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The substrates for the matrix metalloproteases (MMPs) are:

*MMP-1: tissue collagenase: collagen 1, 2, 3, 4, 6, & 10

*MMP-2: gelatinase: gelatins, collagens 4, 5, & 7

*MMP-3: stromleysin-2: casein, fibronectin, laminin, elastin

*MMP-7: matrilysin: casein

*MMP-8: neutrophil collagenase: collagen

*MMP-9: type 4 collagenase: gelatin

*MMP-10: stromelysin-2: casein

*MMP-12: metalloelastase: elastin

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Invitrogen Zymogram gels are an excellent tool for detecting and characterizing proteases that can utilize casein or gelatin as a substrate. Proteases are run on a gel containing one of these substrates under SDS conditions. They can be visualized as clear bands against a dark background following a simple renaturing and staining protocol. These gels are commonly used to detect matrix metalloproteases, a family of enzymes linked to the pathogenesis of cancer and some inflammatory diseases.

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The Zymogram buffers should be stored at 4 degrees C.

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The shelf life of Zymogram gels is 8 weeks when stored at 4 degrees C.

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We recommend storing the Zymogram gels at 4 degrees C.

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Our Zymogram gels do not contain the substrate (gelatin or casein) in the stacking gel although a small amount might mix in during casting of the gel.

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Yes, our Zymogram gels do contain a 4% stacking gel that is ~8 to 9mm long.

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The recommended sample loading volumes and protein loading amounts for the different well formats can be found at: https://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-gel-electrophoresis/protein-gels/recommended-well-loading-volumes-sample-loads.html.

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The ratio of acrylamide:bisacrylamide in our Zymogram gels is 37.5:1 and percentage of cross-linker is 2.6%.

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Our Zymogram gels do not contain SDS. We recommend using the Invitrogen Tris-Glycine SDS Sample buffer and the Invitrogen Tris-Glycine SDS Running buffer with these gels. Invitrogen Zymogram Renaturing buffer and Invitrogen Zymogram Developing buffer are recommended for enzyme assays.

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The Invitrogen 10% Zymogram Gelatin gels are 10% Tris-Glycine gels with 0.1% Type A gelatin from porcine skin (Sigma Cat. No. G2500), as the substrate.

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