Affinity Capture-Western

An interaction is inferred when a bait protein is affinity captured from cell extracts by either polyclonal antibody or epitope tag and the associated interaction partner identified by Western blot with a specific polyclonal antibody or second epitope tag. This category is also used if an interacting protein is visualized directly by dye stain or radioactivity. Note that this differs from any co-purification experiment involving affinity capture in that the co-purification experiment involves at least one extra purification step to get rid of potential contaminating proteins.

Publication

Functional assay of effectors of ADP ribosylation factor 6 during clathrin/AP-2 coat recruitment to membranes.

Krauss M, Haucke V

Arf proteins play pivotal roles in membrane traffic, cell signaling, and actin cytoskeletal rearrangements. We describe here methods to functionally analyze interacting partner proteins of recombinantly produced N-myristoylated Arf6. Combined evidence from affinity purification and chemical crosslinking experiments, in vitro recruitment assays, and the analysis of lipid kinase activities indicates that Arf6-GTP facilitates clathrin/AP-2 recruitment to synaptic membranes by direct ... [more]

Meth. Enzymol. Jan. 18, 2006; 404(0);388-98 [Pubmed: 16413285]

Throughput

  • Low Throughput

Curated By

  • BioGRID