Blue native polyacrylamide gel electrophoresis (BN-PAGE) is a powerful strategy for the separation of native multiprotein complex (MPC). Combined with other techniques, such as SDS–PAGE, immunoblotting, mass spectrometry, and antibody-shift assay, BN-PAGE has been widely used in MPC research. Here, we describe the materials and methods for the basic techniques mentioned above and provide the extensive practical advice and potential pitfalls of BN-PAGE.