The Protein Man's Blog | A Discussion of Protein Research

ImmunoPrecipitation (IP) is a very useful immunochemical technique for isolating and purifying target proteins out of a complex sample (lysate). In conjunction with western blotting and/or other assay techniques, IPs can be used to determine: the presence and quantity of a protein; molecular weight of a polypeptide; rate of synthesis or degradation; enzymatic activity; and to identify certain post-translational modifications and interactions with other proteins, nucleic acids and ligands. There are different IP strategies to purify and analyze your target protein. Read on to see what IP strategy is perfect for you.

While agarose beads have been traditionally used for immunoprecipitation (IP) and other micro-scale protein and antibody purification procedures, magnetic beads have since taken lead and are now considered tool for protein immunoprecipitation.

Commercially available affinity purification supports are designed on the principle of specific surface interactions among biomolecules, including, but not limiting to, antigen-antibody, enzyme-ligand and lectin-carbohydate. Affinity chromatography is one of the most efficient tools used for purification of biomolecules of interest, including proteins, glycoproteins, lipids and nucleic acids. In affinity chromatography, one of the interacting molecules is covalently bound to the resin and is addressed as a ligand. The stationary ligand bound resin interacts with the ligand interacting proteins or biomolecules, which are passed through the resin in mobile phase and hold them to the resin. These molecules are later eluted as a purified fraction with an elution buffer.

Affinity chromatography is the most popular and widely used method for purification of biomolecules including proteins, antibodies, lectins, carbohydrates, nucleic acids and glycoproteins. Affinity purification is based upon specific surface interactions among biological molecules such as antigen-antibody, enzyme-ligand etc. These specific surface interactions enable the purification of molecules from 1000 to 10,000 fold in a single step. In affinity chromatography, the ligand, antigen or one of the interacting molecules is covalently bound to the matrix and is used as a bait to fish out the target protein from any complex pool of proteins including cell or tissue lysate.