Immunoprecipitatie

Immunoprecipitation is used to isolate a single protein from a complex mixture, such as a cell extract.

It employs antibodies specific against the target protein. These antibodies are immobilized on magnetic or agarose beads by direct covalent linking or via antibody-binding recombinant bacterial proteins, such as protein A or G.

When incubated with the protein sample, the pre-coated beads allow the antibodies to form a complex with the target proteins.

Next, the sample is subjected to low-speed centrifugation to precipitate the antibody-protein complex.

The pellet is then resuspended in a mild buffer with low pH or high salt conditions. This breaks the protein-antibody bonds and releases the target protein into the solution.

Finally, the suspension is again centrifuged at a low speed to obtain the target protein in the supernatant.

Like immunoprecipitation, co-immunoprecipitation uses the same principle to study protein-protein interactions.

In this case, a member protein from a protein complex is used as bait to separate its other binding partners.