Antibody Validation for Western Blotting

Western blotting remains one of the most common scientific methods for monitoring protein expression in cells or tissue. The accuracy of western blot results relies heavily of the quality of the primary antibody employed in the immunoblotting. Cell Signaling Technology (CST) provides the highest quality primary and secondary antibodies available for western blotting. CST™ antibodies are produced in-house and validated extensively according to a rigorous protocol.

Validation Steps Include

• Examination of several cell lines and/or tissues of known expression levels allows accurate determination of species cross-reactivity and verifies specificity.
• Treatment of cell lines with growth factors, chemical activators or inhibitors, which induce or inhibit target expression, verifies specificity. Phosphatase treatment confirms phospho-specificity.
• The use of siRNA transfection or knockout cell lines verifies target specificity.
• Side-by-side comparison of lots to ensures lot-to-lot consistency.
• Optimal dilutions and buffers are predetermined, positive and negative cell extracts are specified, and detailed protocols are already optimized, saving valuable time and reagents.