<Li> Co-immunoprecipitation is considered to be the gold standard assay for protein--protein interactions, especially when it is performed with endogenous (not overexpressed and not tagged) proteins . The protein of interest is isolated with a specific antibody . Interaction partners which stick to this protein are subsequently identified by Western blotting . Interactions detected by this approach are considered to be real . However, this method can only verify interactions between suspected interaction partners . Thus, it is not a screening approach . A note of caution also is that immunoprecipitation experiments reveal direct and indirect interactions . Thus, positive results may indicate that two proteins interact directly or may interact via one or more bridging molecules . This could include bridging proteins, nucleic acids (DNA or RNA), or other molecules . </Li> <Li> Bimolecular fluorescence complementation (BiFC) is a new technique in observing the interactions of proteins . Combining with other new techniques, this method can be used to screen protein--protein interactions and their modulators, DERB . </Li> <Li> Affinity electrophoresis as used for estimation of binding constants, as for instance in lectin affinity electrophoresis or characterization of molecules with specific features like glycan content or ligand binding . </Li> <Li> Pull - down assays are a common variation of immunoprecipitation and immunoelectrophoresis and are used identically, although this approach is more amenable to an initial screen for interacting proteins . </Li>

Which one of the following tools is not used to study protein-protein interactions