Additional bands on SDS-PAGE under reducing conditions - (Feb/09/2007 )

We frequently encounter two bands of approximately 60-65 kDa on SDS-PAGE gels (4-12% bis tris from Invitrogen) under reducing conditions (reducing agent from invitrogen as well) and the gels stained with SilverXpress silverstaining from Invitrogen. However, by far not in all gels. Preliminary data seem to indicate that increasing the reducing agent concentration makes it worse. Invitrogen mentions something about "problematic reducing agent and/or the LDS sample buffer" and will sent new aliquots. We don't believe this since we see it with different reducing agents.

Any idea what these bands are and where they come from.

these artifacts have been identified as keratins from dust (dead skin cells). older preparations of reducing agent seem to contain more.

there was a paper where the investigator dipped their finger in the upper reservoir buffer just before running and showed an increase in the artifacts.

the way to limit the amount of artifact seen is to thoroughly clean your apparatus and protect from dust. use the freshest reducing agent you can. and make sure that none of the reagents or buffers touch skin.