Incorrect Antibody Selection (Western Blot (Sigma Protocol))

Question

Accepted Answer

No signal or unexpected banding pattern despite proper protocol execution Common causes: 1) Primary antibody not validated for western blotting application 2) Primary antibody cannot react with protein from the species being studied 3) Secondary antibody cannot bind to primary antibody from its species Solutions: 1) Verify antibody is validated for immunoblotting in product literature 2) Check antibody species reactivity matches your sample species 3) Test secondary antibody by spotting primary antibody on membrane and probing with secondary alone 4) Ensure secondary antibody is raised against the species of primary antibody