 Western blot antibody specificity control

A known target-containing sample is a positive comparison for Western blot antibody detection. A matched sample with gene deletion supplies a negative comparison: disappearance of the proposed band supports the assignment, while a persisting band indicates binding unrelated to the deleted target. Deletion must actually remove the recognized protein; changes in other proteins can complicate the comparison. Omitting the primary antibody checks binding or background from the secondary antibody and detection reagents, but cannot exclude binding of the primary antibody to an unintended protein. Independent antibodies recognizing different epitopes add evidence. ^[ref]