Smeared Bands Troubleshooting in Western Blots

Western Blot / WB is used to detect specific proteins in the given samples. Sometimes, smeared bands may appear in a Western Blot / WB, this makes it difficult for you to figure out which is your protein of interest.

Luckily, we have prepared some possible causes and corresponding solutions for smeared bands in the following Western Blot troubleshooting guide. This guilde can help you solve smeared bands issues and get reliable results in your western blotting.

Western Blot possible causes & solution for smeared bands

Smeared bands in a western blot

Smeared bands in a Western Blot
Possible cause Solution
Particulates in the sample • Remove particulates in the sample by centrifugation prior to loading the gel. Sometimes, using a lithium dodecyl sulfate based loading buffer can give better dissolution of the sample.
Too much sample loaded • Titrate down the amount of protein loaded per lane.
Poorly polymerized gel • To ensure sharp banding, always use fresh APS and TEMED, and allow at least 30 minutes for the gel to polymerize before running.
Gel run too fast • The optimal voltage is mostly determined by the apparatus used. Check with the manufacturer to be sure your run conditions are within the recommended range for that apparatus.