Why am I seeing multiple or unexpected bands on a Western with my affinity purified antibody?

It's common to see multiple bands even when using affinity-purified antibody. This doesn't mean that there's a problem with the antibody’s specificity. Rather, this typically occurs for the following reasons: The native protein shows a different molecular weight than previously predicted. The antibody recognizes either cleaved fragments of the native protein at lower molecular weights, or post-translational modification or aggregated dimers/trimers of the native protein at higher molecular weights. The anti-peptide antibody recognizes a homologous protein in the sample that shares one or more epitopes with the peptide sequence. Unknown protein in lysate sample non-specifically binds to primary antibody or secondary antibody.