Novex™ Tricine Mini Protein Gels, 10%, 1.0 mm

The Tricine gel system, first described by Schagger and von Jagow in 1987, is a modification of the Laemmli Tris-Glycine system to allow for better resolution of smaller proteins and peptides. In the Laemmli system, the proteins are "stacked" in the porous top portion of the gel (stacking gel) between a highly mobile "leading" chloride ion present in the gel buffer and the slower "trailing" glycine ion supplied by the running buffer. These concentrated, thin bands of protein undergo sieving once they reach the resolving gel, which separates them by size.

The resolution of smaller proteins (under 5 kDa) is hindered by the continuous accumulation of free dodecyl-sulfate (DS) ions (from the SDS sample and running buffers) in the stack. This build-up of DS leads to convective mixing of the DS ions with the smaller proteins, causing fuzzy bands and decreased resolution. The mixing of the DS ions with the small proteins will also interfere with the fixing and staining process later. To solve this problem, Schagger and von Jagow replaced the trailing glycine ion with a faster moving Tricine trailing ion. Many small proteins which run with the stacked DS in the Tris Glycine system will separate from DS in the Tricine gel system, resulting in sharper, cleaner bands and better resolution.

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

There may be too much beta-mercaptoethanol (BME), sample buffer salts, or dithiothreitol (DTT) in your samples. If the proteins are over-reduced, they can be negatively charged and actually repel each other across the lanes causing the bands to get narrower as they progress down the gel.

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

Barbell-shaped bands are a result of loading too large a sample volume.

When a large sample volume is loaded, part of the sample tends to diffuse to the sides of the wells. When the run begins and the sample moves through the stacking portion of the gel, the sample will stack incompletely, causing a slight retardation of the portion of the sample that diffused to the sides of the wells.

This effect may be intensified in larger proteins, whose migration is more impeded in the low concentration acrylamide of the stacking gel.

To alleviate the problem, concentrate the protein and load a smaller volume. This gives a "thinner" starting zone.

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

Some potential causes are:

1) Re-oxidation of protein during run

2) Protein has highly hydrophobic regions where protein can exclude SDS.

Steps you can take to improve results:

1) Reduce samples right before loading, and add antioxidant to running buffer. Do not use samples that have been stored in reducing agent.

2) Load sample with 2X sample buffer instead of 1X.

3) Add SDS to upper chamber buffer: try 0.1, 0.2, 0.3, and 0.4% (don't go any higher than 0.4%)

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

Yes. All detergents and even phospholipids in cell extracts will form mixed micelles with SDS and migrate down into the gel.

They can also interfere with the SDS:protein binding equilibrium. Most of the nonionic detergents significantly interfere with SDS-PAGE.

We recommend that you keep the ratio of SDS to lipid or other detergent at 10:1 (or greater) to minimize these effects.

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.