Comparing Co-immunoprecipitation and immunoprecipitation

Posted by beauty33 on January 10th, 2020

What is Immunoprecipitation (IP)

Immunoprecipitation is one of the most widely used methods for antigen detection and purification. The principle of immunoprecipitation (IP) is: an antibody against a specific target protein forms an immune complex with the target protein in the sample (cell lysate, etc.), and then uses the Protein A-magnetic beads or Protein G-magnetic beads to form the immune complex. Finally, the target protein is eluted from the magnetic beads (if the antibody is not covalently attached to the magnetic beads or the denaturing buffer is used to elute the antibody, the antibody will also be eluted), and SDS-PAGE, Western Blot and other means. The eluted protein was analyzed.

What is Co-immunoprecipitation (Co-IP)

Co-IP is an extension of immunoprecipitation and is mainly used for detection of protein-protein interactions. The principle of Co-IP is to capture and purify the target protein based on the IP reaction. If the target protein that interacts with the target protein exists in the sample solution, it will also be captured and purified together, and then obtained by SDS-PAGE, Western Blot and other methods. Analysis of the target protein.

The below is a detailed comparison between IP and Co-IP:

Definition:

1. Immunol precipitation (IP) is a method to purify and enrich the protein of interest by using antibody-specific reactions.

2. Co-immunoprecipitation (Co-IP) is a classic method for studying protein interactions based on the specific interaction between antibodies and antigens. It is an effective method to determine the physiological interaction of two proteins in intact cells method.

Principle:

1. Immunoprecipitation is based on the principle of visible precipitation formed by soluble antigens and corresponding antibodies in the presence of electrolytes at appropriate proportions.

2. The basic principle of co-immunoprecipitation is to add antibody to the protein of interest in the cell lysate, and then add the S. aureus protein A (SPA) that specifically binds to the antibody and binds to Pansobin beads. There is a protein of interest that is bound to the protein of interest, so a complex can be formed-protein of interest-protein of interest-anti-protein of interest-SPA \ | Pansobin, because SPA \ | Pansobin is relatively large, so the complex is centrifuged.

Application:

1. Immunoprecipitation is mainly used for qualitative detection of antigens or antibodies.

2. Co-immunoprecipitation is used to detect and determine the related protein-protein interactions under physiological conditions.

Antigen binding:

1. Immunoprecipitation: A single specific antibody binds to its corresponding antigen, causing it to aggregate and precipitate. That is, immunoprecipitation is to see if there is antigen-antibody binding.

2. Co-immunoprecipitation: Co-immunoprecipitation is the combination of antibody a with antigen A and antibody b with antigen B. If antigen A can interact with and bind to antigen B, aAbB will be precipitated together. Co-immunoprecipitation is to see if the two antigens interact.

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