immunopresipitio

Immunoprecipitation is a laboratory technique used to isolate a specific antigen from a mixture by exploiting the specificity of an antibody that binds to that antigen. In a typical protocol, an antibody is incubated with a biological sample containing the target protein, allowing immune complexes to form. The antibody is captured using a solid support, usually agarose or magnetic beads coated with Protein A or Protein G, and the complexes are collected by centrifugation or magnetic separation. After washing to remove non-specific material, the precipitated proteins are eluted and analyzed by methods such as SDS-PAGE and Western blot, mass spectrometry, or further functional assays.

A related approach is co-immunoprecipitation (co-IP), where the antibody pulls down not only the target protein but also interacting partners, enabling study of protein–protein interactions. Pull-down assays can use recombinant tagged proteins as baits to capture binding proteins from cell lysates. Immunoprecipitation can also be combined with cross-linking to stabilize weak or transient interactions.

Key reagents include high-affinity, specific antibodies; solid supports such as Protein A/G–coupled beads; and appropriate controls, including non-specific IgG and pre-immune serum. Critical steps include optimization of antibody amount, buffer composition, salt concentration, and washing stringency to minimize non-specific binding.

Applications span basic research and diagnostics, including mapping protein interaction networks, purifying protein complexes, validating biomarkers, and enriching proteins for proteomic analyses. Limitations include dependence on antibody quality, potential disruption of interactions during processing, and the risk of artefacts from overexpression or tagging.