Co-immunoprecipitation (Co-IP) Analysis Service


Introduction

Co-immunoprecipitation, Co-IP in short, is  a widely applied technique to identify physiologically-relevant protein-protein  interactions by utilizing target protein-specific antibodies to indirectly  capture proteins that are bound to this specific target protein. As an extension  of IP, Co-IP can capture and purify not only the primary target, but also other  macromolecules binding to the target by native interactions. Difference between  IP and co-IP is the focus of the experiment. IP is focused on the primary  target, which binds the antibody. Whereas, Co-IP targets the secondary targets,  which interacts with the primary proteins, instead of antibody. Following  immunoprecipitation, proteins trapped on the beads are washed and eluted to  gain purified primary and secondary target proteins. These target proteins can  be characterized in various methods, such as WesternBlot and mass spec analysis  under shotgun strategy, to identify partner protein IDs, binding affinities,  kinetics of binding and undiscovered functions of the primary protein.

Co-immunoprecipitation (co-IP)

Critical factors about  Co-IP:

As a powerful technique, Co-IP is utilized  regularly by biochemists to explore protein–protein interactions. While the  Co-IP methodology is straightforward, identifying physiological protein-protein  interactions through Co-IP reaction is not easy, because of the instability of  the interaction, nonspecific binding to IP reagents and antibody contamination.  All of the problems above can have negative effects on detection of  protein-protein interactions.

Co-IP  of protein complexes

Since Co-IP depends on protein-protein  interactions to detect the bound proteins, the binding  affinity and stability of the physiological interactions during the whole Co-IP  process is very important. Inadequate binding time and extensive washing steps  may reduce the binding efficiency and cause a failure of detection of protein  interaction. Therefore, for proteins, which are predicted to have week binding  affinity or dynamic interactions, advance stabilization of protein-protein  interactions are necessary, for instance, crosslinking processes can be  performed before Co-IP.

Unspecific  interactions

Breakage of cell membrane and organelles  cause releasing of large of amount of proteins, which are separated by the  boundary, to come into contact. Therefore, it is inevitable that unspecific  interactions, in other words, false positive interactions, may occurs, which  interfere with the data analysis. This is especially common for proteins that  have unfolded or flexible regions, which are relatively sticky and  unspecifically binds other proteins. Ways to relive such problems can be  preclearance of lysate using primary antibodies to remove unspecific proteins,  and changing the ionic strength of the buffer to reduce the unspecific binding.

Creative  Proteomics can provide custom experimental design  and refine Co-IP parameters based on customers’ needs. We promise reliable  Co-IP analysis of protein-protein interactions with our experienced scientists  and technicians.

Features  of Co-IP service:

Our  Co-IP service contains:

Ordering Procedure:

Ordering Procedure


* For Research Use Only. Not for use in diagnostic procedures.
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