Bands are misshapen or uneven in western blot

Voltage may have been too high during migration.

If the voltage is too high, migration will occur too quickly.

• Check our western blot protocol for the suggested voltage and decrease if necessary.
• Running migration overnight with constant amp at a lower voltage is an option for larger proteins.

Gel may have been too hot during migration.

If the temperature is too high, the pH of the buffer may be slightly altered, which could affect migration.

• Run the gel at 4°C: on ice or in a cold room.
• Consider pre-chilling the running buffer.

The gel has polymerized unevenly.

When the gel has not polymerized properly, bands can appear wonky or uneven. In extreme cases, lanes probed for the same protein can appear at different molecular weights.

• Check your gel recipe to see if you've added the right amount of TEMED.

• Ensure the gel is covered entirely in buffer while it is setting.

The dye has not migrated evenly.

Curving/warping of the dye front can be due to overheating, overloading, field effect, or buffer issues, among others.

• Monitor the dye front for uniform migration.
• Follow recommended guidelines from your apparatus/gel/buffer manufacturer and refer to our protocol.

Wells were overloaded or an uneven amount of protein was loaded.

Too much protein in the well may result in the lane warping.

• Ensure you don't overload wells.
• Refer to the gel manufacturer/comb size recommendations for volumes of lysates to load.

The buffer composition is wrong.

Incompatible sample buffer composition may result in streaking/smearing or uneven gel migration.

• Check for adequate salt/detergent/lysis buffer concentrations for sample preparation.