immunoprecipitated

Immunoprecipitated describes a sample that has been isolated from a complex mixture by means of an antibody that specifically recognizes a target antigen. In this approach, the antibody binds to the antigen, forming an immune complex that can be captured and separated from the rest of the sample, typically using protein A or protein G–containing beads or magnetic beads.

A typical immunoprecipitation workflow involves lysing cells or tissues to release proteins, adding a specific antibody to bind the target, and then using beads to capture the antibody–antigen complexes. The complexes are collected by centrifugation or magnetic separation, followed by multiple washes to remove non-specifically bound material. The immunoprecipitated material can be eluted under denaturing or native conditions for downstream analysis, such as SDS-PAGE and Western blotting, or subjected to mass spectrometry to identify co-precipitating proteins.

Variants of the method include standard immunoprecipitation to enrich a single protein and co-immunoprecipitation (co-IP) to study protein–protein interactions by pulling down one protein along with its binding partners. Reverse immunoprecipitation uses a second antibody to capture a complex from a different angle.

Key considerations for successful immunoprecipitation include antibody specificity and affinity, sample lysis and buffer conditions, and the choice of controls to assess non-specific binding. Common controls include isotype-matched antibodies and no-antibody controls. Immunoprecipitated samples enable targeted analyses while enriching low-abundance proteins from complex mixtures.