what is negative affinity purification?

Affinity purification is a method commonly used in protein chemistry to isolate a product from a mixture that interacts with another known, and previously isolated, product. Negative affinity purification, on the other hand, is a method used to deplete a known contaminant from a protein mixture based upon its ability to bind to a different molecule.

For example, a scientist wants to make an antibody specific for a novel protein. He/she produces the protein as a fusion product linked to GST (glutathione-S-transferase), since there are many methods to allow isolation of this fusion protein (by affinity purification techniques). This fusion product is then injected into a mouse or rabbit and after some time the animal produces antibody in its blood.

While it might be acceptable to use the whole serum (containing the protein specific antibody) from this animal for subsequent experiments, the antibody of interest is only a small fraction of the total protein in the serum. So, the scientist purifies the antibody directed against the protein of interest using a column containing the GST-fusion product used for inoculation (again, affinity purification) and elutes the antibody fraction that recognizes this protein. Great, but the scientist knows that the GST component of the fusion protein will also induce the mouse to make antibodies, and that these GST-specific antibodies will be purified by the previous step. So, he/she decides to do NEGATIVE Affinity Purification by passing the purified antibody mix over a column that contains the GST protein alone (not the fusion product) and then collects the antibody that DOES NOT bind to the GST alone. Voila, protein specific antibody without contaminating GST-specific antibody... using negative affinity purification.