Applications of IP, Co-IP, ChIP, RIP

1. Applications of Immunoprecipitation(IP): to test biochemical properties of proteins

• • Protein purification:

Utilize specific antigen-antibody reaction.

• • Eliminate target protein:

Utilize specific antigen-antibody reaction to eliminate target protein.

• • Optimize western blot:

Enrich target protein (antigen) by IP (up to 10,000 fold enrichment)

Upon enrichment it's easier to get a good western blot result.

• • Detect the modification site of proteins:

Performing mass spectra assay after IP can supply information of modifications of antigens such as phosphorylation, methylation, acetylation and glycosylation.

• • Test the degradation rate of proteins:

Via IP purify the antigens from samples teart ed by actidione. Using western blot to detect contents of antigens in different samples.

Or, firstly label the cells with isotope pulse, then via immunoprecipitation we can measure content of target antigen by autoradiography.

• • Test activities of enzymes:

Purify target enzyme by IP and perform enzyme activity assay.

2. Applications of Co-Immunoprecipitation, Co-IP: to test the proteins' interactions

• • Find a new protein that interacts with a known one:

After typical Co-IP, identify the new protein that binds with a known protein by western-blot or (and) mass spectrometry.

• • Prove a interaction between target protein and tested protein:

After typical Co-IP, perform western blot test with tested protein's antibody to prove its existence. Thus tested protein is proved to interact with known protein.

3. Chromatin immunoprecipitation, ChIP: to test interactions between protein and DNA

• • Test the dynamic interaction:

Purify and amplify the DNA fragments produced by ChIP by PCR and analyze the gene information to get information of dynamic interaction of protein and DNA.

• • Test the relationship between histone modification and DNA expression:

ChIP the target gene and histones using antibodies for different modifications of histones. Evaluate the relationship of histone modification and target gene expression by analysis of interactions of histone modification and promoters of target gene.

• • Test interactions of proteins and RNA:

Process RNA-IP (RIP) developed from ChIP.