Western Blot Troubleshooting Guide

Western Blotting, also known as protein blotting or immunoblotting, is a conventional method to detect specific proteins from a complex mixture of proteins. The basic procedure of Western Blot includes: (1) samples preparation, (2) proteins separated by gel electrophoresis, (3) transfer proteins to a membrane, (4) block nonspecific sites, (5) primary antibody incubation, (6) secondary antibody incubation, (7) detection and Imaging.

However, due to its tedious workflow, every single step of Western Blotting could have gone wrong. Based on 10+ years of experience, we recommend that one should carefully follow your protocols and optimize experimental conditions, this can reduce the possibility of making problems. Additionally, this page lists 10 common problems happened in Western Blotting, and you can easily troubleshoot these problems.

If you have any issues in your Western Blotting, please contact support@sinobiological.com.

Gallery of Western Blot troubleshooting guide

  • High background in a western blot

    High background in a Western Blot means a high signal / noise ratio, and it affects ......

  • Speckled background in a western blot

    Uneven white spots usually appear on a film, making the western blot ......

  • Multiple bands/too many bands  in a western blot

    Multiple bands may increase difficulties and troubles in
    ......

  • No bands in a western blot

    Why are there no bands in a western blot film? This Western Blot ......

  • Low signal/no signal in a western blot

    Low signal in western blotting often means poor transfer quality. What are ......

  • Nonspecific bands in a western blot

    About western blot antibody non-specific binding problem, multiple bands ......

  • Smeared bands in a western blot

    Smeared bands may appear in a Western Blot / WB, this makes it difficult ......

  • Black dots in a western blot

    Black dots will appear in a Western Blotting if antibodies bind to the ......

  • White bands in a western blot

    Why are there white bands in a Western Blot / WB? The following troubleshooting ......

  • Smile effect in a western blot

    Wrong band size and smile effect are also common issues in the Western Blotting ......