Western Blot Application Guide

How western blotting works

Proteins are separated by size using gel electrophoresis, transferred to a membrane and probed with antibodies. Detection may use chemiluminescent, fluorescent or other compatible readouts.

1. Prepare samples using an appropriate lysis and denaturation strategy.
2. Separate proteins by gel electrophoresis.
3. Transfer proteins to a membrane and verify transfer quality.
4. Block, probe, detect and interpret the result with suitable controls.

Experimental planning and controls

Choose an antibody with evidence for western blot in the relevant species and consider expected molecular weight, isoforms and post-translational processing. Include positive and negative sample context where possible. A band at the expected molecular weight alone is not proof of specificity.