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HOME > Learn > Principle and method of the experiment > The principle and method of co-immunoprecipitation (Co-IP)

In Co-IP, complexes of two (or more) proteins are isolated using a procedure similar to the IP procedure.
Co-IP is often used for the analysis of interactions of multiple proteins and their functions.

The principle and method

The principle of Co-IP is the same as IP, except that the proteins associated with the antigen are also precipitated.
Principle of co-immunoprecipitation

A protein complex is isolated by Co-IP using an antibody for one of the components in the complex.
The choice of antibody is critical for successful Co-IP. The antibody must bind to the surface of the complex. Alternatively, a component is tagged, and IP is performed using an anti-tag antibody. In either case, the method of cell lysis and the conditions of washing should be carefully evaluated because components in protein complexes are often held in place by weak interactions.

A common approach is to optimize the condition for Co-IP using a tagged protein, which is easy to detect, and then perform Co-IP experiments with endogenous proteins.
[Related topics] Pull-down assay experiments using Tagged protein purification kits

Complex formation identified by Co-IP should be confirmed by other methods, such as analysis of protein interactions in vivo using fluorescent-labeled proteins.
[Related topics] Protein-Protein Interaction analysis tool "Fluoppi"
[Related topics] Protein-Protein Interaction Detection System "CoralHue™ Fluo-chase Kit"

AZ's point
There are different issues to consider when performing Co-IP with endogenous proteins or tagged recombinant proteins.

Comparison of Co-IP with endogenous proteins versus tagged proteins
Endogenous proteins Tagged proteins (pull-down assay)
Main advantages Protein complexes are isolated in a relatively natural state. An N- or C-terminal tag is likely available for antibody binding after complex formation. Antibody binding is unlikely to interfere with complex formation.
Issues to consider The epitope may be buried upon complex formation. Antibody binding may interfere with complex formation. The expression levels of recombinant proteins are substantially higher than those of their endogenous counterparts, which may result in artifactual results.

[Related topics] Links to Co-IP-related products
Anti-tag antibodies suitable for IP List of monoclonal anti-tag antibodies
List of polyclonal anti-tag antibodies
List of anti-tag antibody-coated agarose beads
List of anti-tag antibody-coated magnetic beads
List of anti-tag antibody-coated magnetic agarose beads
Pull-down assay-related kits,
gels and elution peptides
List of tagged protein purification kits
List of tagged protein purification gel and elution peptide sets
Primary antibodies suitable for IP (excluding anti-tag antibodies) List of monoclonal primary antibodies
List of polyclonal primary antibodies
List of antibody-coated agarose beads
List of antibody-coated magnetic beads
List of antibody-coated magnetic agarose beads
Others Protein G-coated magnetic beads
Magnetic racks
For analysis of immunoprecipitated proteins HRP-DirecT series

Co-eluted antibody is not detected with HRP-labeled antibodies in Western blotting of immunoprecipitated proteins. It is especially useful when the target protein band overlaps with the heavy or light chain band. Analysis time is also reduced.
For confirmation of complex formation Protein-Protein Interaction analysis tool "Fluoppi"
Protein-Protein Interaction Detection System "CoralHue™ Fluo-chase Kit"

Related links

Fractionation and purification of proteins